New transition metal ion complexes with benzimidazole-5-carboxylic acid hydrazides with antitumor activity

Article abstract of DOI:10.1016/j.ejmech.2008.07.013

Metal complexes of 2-methyl-1H-benzimidazole-5-carboxylic acid hydrazide (4a; L₁) and its Schiff base 2-methyl-N-(propan-2-ylidene)-1H-benzimidazole-5-carbohydrazide (5a; L₂) with transition metal ions e.g., copper, silver, nickel, i

Full text of DOI:10.1016/j.ejmech.2008.07.013

European Journal of Medicinal Chemistry 44 (2009) 1500–1508
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
New transition metal ion complexes with benzimidazole-5-carboxylic acid
hydrazides with antitumor activity
,
b
a
c
c
Shadia A. Galal a , Khaled H. Hegab , Ahmed S. Kassab , Mireya L. Rodriguez , Sean M. Kerwin ,
*
Abdel-Mo’men A. El-Khamry d, Hoda I. El Diwani a
a Division of Pharmaceutical and Drug Industries, Department of Chemistry of Natural and Microbial Products, National Research Center, Cairo, Egypt
b Division of Inorganic Chemical Industries and Mineral Resources, Department of Inorganic Chemistry, National Research Center, Cairo, Egypt
c Division of Medicinal Chemistry, Department of Chemistry and Biochemistry, The University of Texas, Austin, TX 78712, USA
d Faculty of Science, Department of Chemistry, Ein Shams University, Cairo, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 1 June 2008
Received in revised form 8 July 2008
Accepted 11 July 2008
Available online 19 July 2008
Metal complexes of 2-methyl-1H-benzimidazole-5-carboxylic acid hydrazide (4a; L1) and its Schiff base
2-methyl-N-(propan-2-ylidene)-1H-benzimidazole-5-carbohydrazide (5a; L2) with transition metal ions
e.g., copper, silver, nickel, iron and manganese were prepared. The complexes formed were 1:1 or 1:2
M:L complexes and have the structural formulae [Cu(L1)Cl(H2O)]Cl$3H2O (6), [Ag(L1)NO3(H2O)] (7),
[Ni(L1)Cl2(H2O)2]$H2O (8), [Fe(L1)Cl3(H2O)]$3H2O (9) and [Mn(L1)2Cl(H2O)]Cl$3H2O (10) for ligand L1, and
[Cu(L2)Cl2(H2O)2]$H2O (11), [Ag(L2)2]NO3$H2O (12), [Ni(L2)2Cl2]$5H2O (13), [Fe(L2)2Cl2]Cl$2H2O (14) and
[Mn(L2)Cl2(H2O)2]$H2O (15) for ligand L2. The antitumor activity of the synthesized compounds has been
Keywords:
2-Methylbenzimidazole
Transition metal complexes
Antitumor activity
studied. The silver complex 7 was found to display cytotoxicity (IC50 ¼ 2
mM) against both human lung
cancer cell line A549 and human breast cancer cell line MCF-7.
Ó 2008 Elsevier Masson SAS. All rights reserved.
1. Introduction
MG-22A (mouse hepatoma), HT-1080 (human fibrosarcoma), B16
(mouse melanoma, Neuro 2A (mouse neuroblastoma)) and normal
mouse fibroblast cells [3].
Benzimidazole ring acts as ligand toward transition metal ions
in a variety of biologically important molecules [1].
Cu(II) complex of CuL(ClO4)2 (L ¼ N,N,N0,N0-tetrakis[2-
benzimidazolyl)methyl]-1,3-diaminopropane (I) can electrostati-
cally bind to the phosphate groups of DNA backbone and partially
intercalate into the double helix of DNA [2].
It was reported that some of the 2-substituted benzimid-
azole Pt(II) complexes have in vitro cytotoxic activities on
different cancer cell lines. cis-[Pt (R(ꢀ) and S(þ)-HBB)2Cl2]
(HBB ¼ 2-
a-hydroxybenzylbenzimidazole) III and IV, respec-
tively, were synthesized and have in vitro cytotoxic activities on
the human MCF-7 breast cancer and HeLa cervix cancer cell
lines [4–8].
N-Trimethylsilylpropylbenzimidazole (II) metal complexes
exhibit cytotoxic activity on four monolayer tumor cell lines:
* Corresponding author. Tel.: þ20 233371718; fax: þ20 233370931.
E-mail address: sh12galal@yahoo.com (S.A. Galal).
0223-5234/$ – see front matter Ó 2008 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2008.07.013

S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
1501
2. Results and discussion
[Mn(L1)2Cl(H2O)]Cl$3H2O (10) for ligand 4a; L1 (cf. Scheme 2). On
the other hand, [Cu(L2)Cl2(H2O)2]$H2O (11), [Ag(L2)2]NO3$H2O (12),
2.1. Chemistry
[Ni(L2)2Cl2]$5H2O
[Mn(L2)Cl2(H2O)2]$H2O (15) for ligand 5a; L2 (cf. Scheme 3).
The molar conductance values of the prepared metal complexes
7–9, 11, 13 and 15 range from 12.3 to 28.9
cm2 molꢀ1, which
(13),
[Fe(L2)2Cl2]Cl$2H2O
(14)
and
2-Methyl-1H-benzimidazole-5-carboxylic acid (1) was prepared
from 3,4-diaminobenzoic acid with acetic acid in hydrochloric acid
according to Philips method [9,10]. Esterification of 1 was per-
formed with ethanol and sulfuric acid to obtain the ester 2 which
was nitrated to yield the 6(5)-nitro derivative 3. The absence of the
meta coupling between the two aromatic protons besides the
presence of two singlets in the 1H NMR spectrum gave the evidence
of the presence of the nitro group in the 5 or 6 position.
U
indicated their non-ionic nature [13]. Complexes 6, 10, 12 and 14
behaved as ionic compounds which gave molar conductance 67, 78,
65 and 76
U
cm2 molꢀ1, respectively.
The results obtained from thermo-gravimetric analyses [14,15]
were in agreement with the suggested theoretical formulae from
the elemental analyses. For example, for complexes 7 and 10, the
mass loss in the temperature range 60–145 ꢁC may be due to the
loss of hydrated water in complex 10, while the mass loss in
the temperature range from 160 to 244 ꢁC, 145 to 271 ꢁC may be
due to the loss of coordinated water molecules in 7 and 10
complexes, respectively.
The two complexes under investigation have two decomposi-
tion steps: the first step for 7 and 10 was at 244–401 ꢁC, 271–
756 ꢁC, respectively, to form unstable intermediates, then finally
decomposition in the range 401–1000 ꢁC, 756–1050 ꢁC, respec-
tively, took place to form Ag and Mn2O3. The amounts of residue
were in agreement with the calculated values.
The acid hydrazides 4(a, b) were formed by the reaction of 2 or 3
with hydrazine hydrate [11]. Compound 4a was condensed with
acetone to afford the hydrazone 5a while 4b condensed with
acetone,
furan-2-carboxaldehyde,
5-methylfuran-2-carbox-
aldehyde, pyridine-2-carboxaldehyde, indole-3-carboxaldehyde
and 4-methylbenzaldehyde to afford 5(b–g), respectively (cf.
Scheme 1).
The metal complexes 6–10 of the hydrazide 4a and the metal
complexes 11–15 of the Schiff base 5a with CuCl2, AgNO3, NiCl2,
FeCl3 and MnCl2, respectively, were synthesized according to the
´
reported methods by Merchan et al. [12] to form 1:1 or 1:2 (met-
al:ligand) complexes. These complexes were characterized by
elemental analyses, IR, 1H NMR (for diamagnetic complexes),
electronic and mass spectra, magnetic moments and conductance
measurements as well as thermo-gravimetric analyses. Based upon
this characterization, the prepared complexes have the structural
formulae: [Cu(L1)Cl(H2O)]Cl$3H2O (6), [Ag(L1)NO3(H2O)] (7),
[Ni(L1)Cl2(H2O)2]$H2O (8), [Fe(L1)Cl3(H2O)]$3H2O (9) and
For complexes 11–13 the mass loss in the temperature range 70–
120 ꢁC, 70–110 ꢁC and 70–135 ꢁC may be due to the loss of hydrated
water in complexes 11–13, respectively, while at 150–280 ꢁC the
mass loss may be due to coordinated water in complex 11.
Also, the first decomposition took place at 292–439 ꢁC, 208–
387 ꢁC and 290–439 ꢁC to form unstable intermediates for 11–13,
respectively, finally, the metal residues CuO, Ag2O and NiO were
NH2
HOOC
HOOC
N
HCl
C2H5OH /
H2SO4
CH3
EtO
CH3COOH
+
N
H
NH2
1
O
O
H2N
N
NH2NH2
N
H
N
CH3
N
H
CH3
R
N
H
4a, R= H
4b, R=NO2
2
R2
R1
N
O
O
R
R1
R2
N
N
H
CH3
N
H
5a-g
5a, R = H, R1 = R2 = CH3
5b, R = NO2, R1 = R2 = CH3
5c, R = NO2, R1 = H, R2 = 2-furyl
5d, R = NO2, R1 = H, R2 = 5-methyl-2-furyl
5e, R = NO2, R1 = H, R2 = 2-pyridyl
5f, R = NO2, R1 = H, R2 = 3-indolyl
5g, R = NO2, R1 = H, R2 = 4-methylphenyl
Scheme 1.

1502
S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
O
H2N
N
N
N
H
CH3
4a
CuCl2
H
AgNO3
O3N
OH2
Cl
OH2
Cu
Ag
H2N
H2N
O
O
N
N
H
H
Cl.3H2O
N
N
CH3
CH3
N
N
H
H
7
6
NiCl2
FeCl3
Cl
OH2
Cl
Fe
Cl
OH2
O
Ni
Cl
OH2
Cl
H2N
H2N
.H2O
O
N
H
3H2O
N
N
N
N
N
H
CH
3
CH3
H
H
8
9
MnCl2
H
N
H3C
N
H
N
OH2
NH2
O
Mn
O
H2N
Cl.3H2O
Cl
N
N
CH3
H
N
H
10
Scheme 2.
formed at 439–637 ꢁC, 387–580 ꢁC and 439–880 ꢁC, respectively.
The amounts of residue were in agreement with the calculated
values (cf. Table 1).
exhibited new bands in all the spectra due to the presence of M–N
bands and M–O bands for 11–15, metal ion complexes, respectively.
Ligand 5a acted as a bidentate ligand.
The IR spectra of complexes 6–15 showed a broad band in the
The analyses by mass spectrometry at 20 eV for compounds 6, 7,
range of 3587–3386 cmꢀ1, due to
n
(OH), suggesting the presence of
water molecules. The retention of broad feature at 2800–
2600 cmꢀ1 and only slight shift of
(C]N) (ring) in these complexes
9, 12 and 14 showed the presence of molecular ion peaks Mþ þ 1,
M
þ ꢀ 2, Mþ,
M
þ þ 2 and
M
þ þ 2, respectively. This behavior
n
confirmed that both ligands are strongly attached to metal ion. In
compound 14 the water molecule was lost and peak at Mþ ꢀ H2O
was observed. Compounds 8, 10, 11 and 15 decomposed without
melting and their mass spectra were not recorded.
suggested the involvement of –NH nitrogen (ring) in hydrogen
bonding and non-involvement of tertiary nitrogen (ring) in coor-
dination [16–19].
The IR spectra of complexes 6–10 showed weakness of broad
The presence of molecular ion peaks was consistent with IR
spectra of complexes which were in agreement with a strong
bonding from ligands toward the metal ions.
bands of
indicated the involvement of carbonyl oxygen in coordination.
Furthermore the presence of (M–N) and (M–O) in the FT-IR
n(NH2) and considerable lower frequency shift of n(C]O)
n
n
The 1H NMR spectra of the diamagnetic complexes 7 and 12
spectra of these complexes indicated the bidentate nature of the
exhibited the presence of only a slight shift of d (CH3 of benz-
ligand 4a. For complexes 11–15, the IR spectra exhibited a consid-
erable lower frequency shift of both n(C]O) and n(C]N) indicating
the involvement of carbonyl oxygen and azomethine nitrogen in
coordination [20–22]. The FT-IR spectra of all the complexes
imidazole) compared to free ligands in these complexes. The
presence of exchangeable proton (NH of ring) signal was in
agreement with IR spectra of complexes where imidazole ring
wasn’t involved in complex formation. The downfield shift of

S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
1503
O
H3C
N
N
N
N
H
CH3
H3C
H
5a
CuCl2
AgNO3
H
N
H
N
CH3
Cl
H3C
H3C
OH2
OH2
N
Cl
N
Cu
O
O
.H2O
Ag
N
N
N
H3C
H3C
CH3
NO3.H2O
N
O
CH3
CH3
H
N
H
N
H
N
H3C
11
N
H
12
FeCl3
NiCl2
H
N
CH3
H
N
N
H
H
N
N
CH3
H3C
H3C
H3C
H3C
N
Cl
N
O
N
Cl
Fe
Cl
N
O
N
Ni
Cl
.5H2O
Cl. 2H2O
O
CH3
CH3
CH3
CH3
O
N
N
N
H3C
N
H3C
H
N
H
H
N
H
13
14
MnCl2
Cl
OH2
OH2
Cl
O
Mn
N
N
CH3
.H2O
H3C
N
N
H
H
H3C
15
Scheme 3.
Table 1
Thermo-gravimetric analyses’ results of the investigated metal complexes 6–15 of ligands 4a and 5a
Compound Structural formula M:L Water elimination
Decomposition Residue (%) (metal oxide or
stages (ꢁC)
metal)
Temperature Hydrated water mass loss % Temperature Coordinated water mass
Temperature Calcd
(ꢁC)
Calcd (Found)
(ꢁC)
loss % Calcd (Found)
(ꢁC)
(Found)
7
Ag(L1)NO3(H2O)
1:1
–
–
160–244
4.77 (3.68)
244–401,
401–1000
271–756,
756–1050
292–439,
439–637
208–387,
387–580
290–439,
439–880
1000
28.54 (28.00)
27.36 (28.12)
10.99 (10.62)
35.76 (35.97)
11.01 (10.93)
10
11
12
13
Mn(L1)2Cl(H2O)Cl$3H2O 1:2 60–145
[Cu(L2)Cl2(H2O)2]$H2O 1:1 70–120
9.36 (9.68)
4.32 (4.50)
2.78 (2.69)
13.27 (13.00)
145–271
3.12 (3.68)
1050
637
580
880
150–280
8.63 (8.82)
[Ag(L2)2]NO3$H2O
[Ni(L2)2Cl2]$5H2O
1:2 70–110
1:2 70–135
–
–
–
–

1504
S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
d
(NH2 of hydrazide of complex 7) compared to free ligand was
[Ag(L1)NO3(H2O)] (7), [Ni(L1)Cl2(H2O)2]$H2O (8), also with
consistent with IR spectra.
[Fe(L1)Cl3(H2O)]$3H2O (9) and [Mn(L1)2Cl(H2O)]Cl$3H2O (10) (1:2
M:L). Also, the ligand 5a behaved as a bidentate ligand in its 1:1 M:L
complexes [Cu(L2)Cl2(H2O)2]$H2O (11) and [Mn(L2)Cl2(H2O)2]$H2O
(15), respectively, but behaved as bidentate ligand in its 1:2 M:L
complexes [Ag(L2)2]NO3$H2O (12), [Ni(L2)2Cl2]$5H2O (13) and
[Fe(L2)2Cl2]Cl$2H2O (14), respectively. The molar conductance
showed the ionic nature of 6, 10, 12 and 14.
Electronic spectra of 8–11, 13–15 were in agreement with the
octahedral environment around the metal ion, but 7 and 12 were in
favor of the tetrahedral one. Moreover, the spectral data together
with the magnetic moment showed a square-planar geometry of
complex 6.
Of the complexes examined, the Ag(I) complex 7 displayed the
most promising anticancer activity against two human cancer cell
lines. The other complexes derived from ligands 4a and 5a were not
cytotoxic, besides the limited solubility of these complexes limited
their evaluation.
The electronic spectrum of complex 4a exhibited bands at 277,
325 nm and that of complex 5a showed bands at 260, 280 nm which
were assigned to
p /
p* and n–p* transitions. Complex 6 had two
strong bands at 277, 325 nm assignable to metal–ligand charge
transfer (MLCT) and only one band at 790 nm (d–d transition). The
magnetic moment at 1.87 B.M. is characteristic of typical square-
planar geometry [23]. The diamagnetic complexes such as 7 and 12
displayedstrongbandsat260, 320 nmand270, 285 nm, respectively,
indicating a tetrahedral geometry of these complexes. Complexes of
4a and 5a showed charge transfer bands at range 255–325 nm.
Complexes 11 and 14 displayed bands at 350, 345 nm, respectively.
Also, complex 8 displayed three bands at 330, 422 and 830 nm
assignable to d–d transitions. Complexes 10, 13 and 15 did not show
the characteristic band in the d–d region. All complexes have
magnetic moments in the range 1.68–1.93 B.M. (low spin complexes)
characteristic of the octahedral geometry [24] (cf. Table 2).
2.2. Cytotoxicity
4. Experimental
The cytotoxicity of the synthetic intermediates 1–3, the ligands
4a–b and 5a–g, and the metal ion complexes 6–15 was tested using
the AlamarBlue assay in breast cancer cell line MCF-7 and lung
cancer cell line A549. For comparison UK-1 and Mitomycin C were
also tested (cf. Table 3). All of the synthetic intermediates and
ligands were non-cytotoxic at the highest concentration examined
4.1. Physical measurements
Microanalyses of the ligands and complexes were performed in
the Microanalytical Laboratory Center, Faculty of Science, Cairo
University, Egypt. Molar conductance was measured at room
temperature on electronic conductivity model 19000-05 (USA). The
IR spectra (4000–400 cmꢀ1) were recorded using KBr pellets in
a Jasco FT/IR 300 E Fourier transform infrared spectrophotometer
and in the 500–100 cmꢀ1 region using polyethylene-sandwiched
Nujol mulls on a Perkin Elmer FT-IR 1650 (spectrophotometer).
The 1H and 13C NMR spectra were recorded using Joel EX-200,
270 and 500 MHz NMR spectrophotometers. The electronic
absorption spectra were recorded using a Shimadzu UV-240 UV–
visible recording spectrophotometer. The mass spectroscopy was
carried out using Finnigan mat SSQ 7000 (Thermo. Inst. Sys. Inc.,
USA) spectrophotometer at 70 and 20 eV. Thermo-gravimetric
analyses were carried out using a DTA-7 and TGA-7 Perkin Elmer 7
series thermal analyses system. The magnetic moments were
measured using Gouy method with Hg[Co(SCN)4] as calibrant.
(100
hydrazone ligand 5g, which has an IC50 of 73
also cytotoxic against MCF-7 cells (IC50 ¼ 3
(IC50 ¼ 81 M) and ethyl ester 2 (IC50 ¼ 62 M). The only metal ion
complex examined here that displayed cytotoxicity was the silver
complex 7, with IC50 values of 2 M against both A549 and MCF-7
m
M) against A549 cell line, with the exception of the acyl
M. Ligand 5g was
M), as were 5c
m
m
m
m
m
cell lines. However, a number of these metal ion complexes derived
from ligand 5a (e.g., 12–15) were not sufficiently soluble in either
DMSO or water to perform cytotoxicity assays. Other complexes
could only be evaluated at concentrations less than 25
(complex 9), 20 M (complexes 6 and 10), or 7 M (complex 11),
due to limited solubility.
mM
m
m
The pronounced cytotoxicity of silver complex 7, relative to the
free ligand 4a and the other metal ion complexes derived from 4a
was striking. While a variety of Ag(I) complexes have been identi-
fied as potential anticancer agents [25], compound 7 was the first
example of a hydrazide-derived Ag(I) complex with cytotoxic
activity in human cancer cell lines.
4.1.1. Preparation of 2-methyl-1H-benzimidazole-5-carboxylic acid
(1) [9,10]
To a solution of 3,4-diaminobenzoic acid (40 mmol) in concen-
trated hydrochloric acid (30 mL) was added glacial acetic acid
(60 mmol). The reaction mixture was heated under reflux for 4 h,
diluted with water (500 mL) and brought to pH 4.5 with aq.
ammonia where the precipitate was formed, it was filtered off and
crystallized from ethanol/water as a brown powder. Rf ¼ 0.43
(chloroform/ethyl acetate/methanol, 1:1:1/5). Yield: 85%, m.p.
3. Conclusion
IR spectra showed that the ligand 4a behaved as a bidentate
ligand in its 1:1 M:L complexes with [Cu(L1)Cl(H2O)]Cl$3H2O (6),
Table 2
Electronic absorption spectral bands and magnetic moment of the ligands 4a, 5a and their complexes 6–15
Compound
Ligand and its complexes
Electronic absorption bands, lmax (nm)
Magnetic moment B/M
Mode
Intraligand and charge transfer bands
d–d Bands
4a
5a
6
7
8
L1 ¼ C9H10N4O
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
277, 325
260, 280
275, 320
260, 275, 320
255, 277
260, 277, 325
260, 285, 325
270, 280, 285
270, 285
265, 285
260, 290
–
–
790
–
330, 422, 830
–
–
350
–
–
–
L2 ¼ C12H14N4O
[Cu(L1)Cl(H2O)]Cl$3H2O
Ag(L1)NO3(H2O)
1.87
Diamagnetic
2.88
1.68
1.93
1.93
Diamagnetic
2.87
1.63
1.98
[Ni(L1)Cl2(H2O)2]$H2O
[Fe(L1)Cl3(H2O)]$3H2O
[Mn(L1)2Cl(H2O)]Cl$3H2O
[Cu(L2)Cl2(H2O)2]$H2O
[Ag(L2)2]NO3$H2O
[Ni(L2)2Cl2]$5H2O
[Fe(L2)2Cl2]Cl$2H2O
[Mn(L2)Cl2(H2O)2]$H2O
9
10
11
12
13
14
15
–
345
–
260, 280, 285

S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
1505
Table 3
(chloroform/ethyl acetate/methanol, 1:1:1/5). Yield: 98%, m.p. 100–
102 ꢁC. 1H NMR (270 MHz, DMSO-d6)
Cytotoxicity of the synthetic intermediates 1–3, the ligands 4a–b and 5a–g, and the
metal ion complexes 6–15 against MCF-7 and A549 cell lines
d
1.35 (t, J ¼ 6.92 Hz, 3H, CH3
ester), 2.8 (s, 3H, CH3 benzimidazole), 4.35 (q, J ¼ 6.92 Hz, 2H, CH2
Compound
IC50 for MCF-7a
(m
M)
IC50 for A549a
(mM)
ester), 8.15 (s, 1H, H4), 8.5 (s, 1H, H7), 9.45 (br, NH, D2O
exchangeable). 13C NMR (270 MHz, DMSO-d6)
d 13.278 (CH3 ester),
1
2
3
4a
4b
5a
5b
5c
5d
5e
5f
5g
6
7
8
9
>100
62 ꢂ 18
>100
>100
>100
>100
>100
81 ꢂ 12
>100
>100
>100
3 ꢂ 0.3
>20b
2 ꢂ 0.3
>100
>25c
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
73 ꢂ 20
>20b
2 ꢂ 1
>100
>25c
>20b
>7d
13.861 (CH3 benzimidazole), 62.720 (CH2 ester), 111.546, 115.831,
123.941, 132.767, 133.843 and 145.339 (Ar–C), 157.722 (C]N),
164.537 (carboxylic ester C]O). IR (cmꢀ1): 3453.33 (NH), 1736.92
(C]O), 2800–2600 (intermolecular H-bonded NH), 1629.96 (C]N),
1545.96 and 1383.28 (NO2). MS: m/z 249 (Mþ, 10.05%). Anal. Calcd
for C11H11N3O4: C, 53.01; H, 4.45; N, 16.86. Found: C, 53.13; H, 4.39;
N, 16.90.
4.1.4. Preparation of 2-methyl-1H-benzimidazole-5-
carbohydrazide (4a) and 2-methyl-6-nitro-1H-benzimidazole-5-
carbohydrazide (4b)
General procedure. Compound 2 or 3 (7 mmol) and hydrazine
monohydrate 98% (10 mL) were stirred at room temperature for
15 h. The reaction mixture was then concentrated under reduced
pressure and triturated with methanol several times to get rid of
the excess hydrazine hydrate, the solid was recrystallized from
ethanol as a brown powder.
10
11
12
13
14
15
>20b
>7d
nde
nde
nde
nde
nde
nde
nde
nde
UK-1
Mitomycin C
1.4 ꢂ 0.9
2.0 ꢂ 0.5
0.2 ꢂ 0.07
0.4 ꢂ 0.1
4.1.4.1. Compound 4a. Rf ¼ 0.44 (petroleum ether/methanol, 2:3).
Yield: 80%, m.p. 260–262 ꢁC. 1H NMR (500 MHz, DMSO-d6)
d 2.45 (s,
a
Determined using the AlamarBlue assay. See Section 4.2 for details.
Highest concentration tested was 20
Highest concentration tested was 25
Highest concentration tested was 7
Not determined due to limited solubility in DMSO or aqueous solutions.
b
c
m
m
mM.
M.
M.
3H, CH3), 4.4 (br, NH2, D2O exchangeable), 7.41 (d, J ¼ 8.5 Hz, 1H,
H7), 7.59 (d, J ¼ 8.5 Hz, 1H, H6), 7.91 (s, 1H, H4), 9.63 (s, 1H, NH
hydrazide, D2O exchangeable), 10.32 (br, NH benzimidazole, D2O
exchangeable). IR (cmꢀ1): 3410.87 (NH benzimidazole), 3331.11,
3296.87 and 3265 (NH2, NH), 2800–2600 (intermolecular
H-bonded NH), 1666.00 (C]O), 1625.62 (C]N benzimidazole). MS:
m/z 190 (Mþ, 100%). Anal. Calcd for C9H10N4O: C, 56.83; H, 5.30; N,
29.46. Found: C, 56.75; H, 5.20; N, 29.35.
d
e
>300 ꢁC. 1H NMR (270 MHz, DMSO-d6)
d 2.5 (s, 3H, CH3), 7.5 (d,
J ¼ 8.5 Hz,1H, H7), 7.8 (d, J ¼ 8.5 Hz,1H, H6), 8.1 (s,1H, H4). 13C NMR
(DMSO-d6)
d 14.74 (CH3), 113.766, 116.372, 122.895, 123.883,
138.797 and 141.978 (Ar–C), 154.11 (C]N), 168.12 (carboxylic
C]O). IR (cmꢀ1): 3425.13 (NH), 3200.51–2400.35 (intermolecular
H-bonded OH), 1700.02 (carboxylic C]O), 1624.12 (C]N). MS: m/z
176 (Mþ, 100%). Anal. Calcd for C9H8N2O2: C, 61.36; H, 4.58; N,15.90.
Found: C, 61.30; H, 4.52; N, 15.86.
4.1.4.2. Compound 4b. Rf ¼ 0.48 (petroleum ether/methanol, 2:3).
Yield: 86%, m.p. 188–190 ꢁC. 1H NMR (270 MHz, DMSO-d6)
d 2.49 (s,
3H, CH3), 4.46 (s, NH2 hydrazide, D2O exchangeable), 7.53 (s, 1H,
H4), 8.16 (s, 1H, H7), 8.95 (br, NH benzimidazole, D2O exchange-
able), 9.70 (s, 1H, NH hydrazide, D2O exchangeable). IR (cmꢀ1):
3406.2 (NH benzimidazole), 3325.4–3265.6 (NH2, NH), 1677.2
(C]O), 1626.7 (C]N benzimidazole), 1524.2 and 1331.2 (NO2). MS:
m/z 235 (Mþ,12.25%). Anal. Calcd for C9H9N5O3: C, 45.96; H, 3.86; N,
29.78. Found: C, 45.83; H, 3.89; N, 29.74.
4.1.2. Preparation of ethyl 2-methyl-1H-benzimidazole-5-
carboxylate (2)
Compound 1 (8 mmol) in absolute ethanol (50 mL) and few
drops of concentrated sulfuric acid were refluxed for 20 h, diluted
with water (500 mL) and brought to pH 7 with NaHCO3 solution
(5%). The precipitate formed was filtered off and crystallized from
ethanol/water as a buff powder. Rf ¼ 0.76 (petroleum ether/ethyl
acetate/methanol, 1:1:1/4). Yield: 79%, m.p. 170–172 ꢁC. 1H NMR
4.1.5. Preparation of compounds 5a–g
General procedure. The metal complexes of the studied ligands
were prepared by mixing a hot ethanolic solution containing the
transition metal ion with the required amount of an ethanolic
solution of the corresponding ligands to form 1:1, 1:2 M:L
complexes. The reaction mixture was stirred at 50 ꢁC, for a time
depending on the transition metal salt used. The resulting precip-
itates were filtered off, washed with diethyl ether followed by
warm ethanol and then dried in a vacuum desiccator over anhy-
drous CaCl2.
(270 MHz, DMSO-d6)
d
1.35 (t, J ¼ 6.92 Hz, 3H, CH3 ester), 2.55 (s,
3H, CH3 benzimidazole), 4.30 (q, J ¼ 6.92, 2H, CH2 ester), 7.5 (d,
J ¼ 8.5 Hz, 1H, H7), 7.75 (d, J ¼ 8.5 Hz, 1H, H6), 8.05 (s, 1H, H4), 12.5
(br, NH, D2O exchangeable). IR (cmꢀ1): 3468.75 (NH), 2800–2600
(intermolecular H-bonded NH), 1712.94 (C]O), 1626.2 (C]N). MS:
m/z 204.1 (Mþ, 62%). Anal. Calcd for C11H12N2O2: C, 64.69; H, 5.92;
N, 13.72. Found: C, 64.77; H, 5.88; N, 13.78.
4.1.3. Preparation of ethyl 2-methyl-6-nitro-1H-benzimidazole-5-
carboxylate (3)
4.1.6. 2-Methyl-N-(propan-2-ylidene)-1H-benzimidazole-5-
carbohydrazide (5a)
In an ice/NaCl bath, concentrated sulfuric acid (2 mL) was added
to compound 2 (5 mmol). The solution was stirred for 30 min, then
a mixture of concentrated nitric acid (0.4 mL) and concentrated
sulfuric acid (0.6 mL) was added dropwise to this solution. The
reaction mixture was stirred for 20 min. The temperature of
the reaction was kept within 0–5 ꢁC range during the reaction. The
reaction then was brought to pH 6–7 with aqueous ammonia at
room temperature. The precipitate formed was filtered off and
crystallized from ethyl acetate as a yellow powder. Rf ¼ 0.78
Buff solid. Rf ¼ 0.47 (ethyl acetate/methanol, 4:1/4). Yield: 91%,
m.p. 203–205 ꢁC. 1H NMR (200 MHz, DMSO-d6)
d 1.97 (s, 3H, CH3
isopropylidine), 2.01 (s, 3H, CH3 isopropylidine), 2.52 (s, 3H, CH3
benzimidazole), 7.30 (s, 1H, NH hydrazide, D2O exchangeable), 7.49
(d, J ¼ 8.5 Hz, 1H, H7), 7.66 (d, J ¼ 8.5 Hz, 1H, H6), 7.99 (s, 1H, H4),
10.28 (br, NH benzimidazole, D2O exchangeable). IR (cmꢀ1):
3410.72 (NH benzimidazole), 3308.73 (NH hydrazide), 2800–2600
(intermolecular H-bonded NH), 1659.99 (C]O), 1627.82 (C]N
benzimidazole), 1619.00 (C]N Schiff base). MS: m/z 230 (Mþ, 9%).

1506
S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
Anal. Calcd for C12H14N4O: C, 62.59; H, 6.13; N, 24.33. Found: C,
62.55; H, 6.18; N, 24.27.
benzimidazole), 12.14 and 13.05 (br, NH benzimidazole, indole, D2O
exchangeable). IR (cmꢀ1): 3489.37, 3431.38, 3359.37 (NH benz-
imidazole, indole, hydrazide), 1660.44 (C]O), 1640.62 (C]N Schiff
base), 1625 (C]N), 1519.53 and 1334.28 (NO2). MS: m/z 362 (Mþ,
18%). Anal. Calcd for C18H14N6O3: C, 59.67; H, 3.89; N, 23.19. Found:
C, 59.70; H, 3.99; N, 23.10.
4.1.7. 2-Methyl-6-nitro-N-(propan-2-ylidene)-1H-benzimidazole-
5-carbohydrazide (5b)
Buff solid. Rf ¼ 0.49 (ethyl acetate/methanol, 4:1/4). Yield: 92%,
m.p. 289–290 ꢁC. 1H NMR (500 MHz, DMSO-d6)
d 1.85 (s, 3H, CH3),
1.90 (s, 3H, CH3), 2.55 (s, 3H, CH3 benzimidazole), 7.30 (s, 1H, NH
hydrazide, D2O exchangeable), 7.51 (s, 1H, H4), 8.19 (s, 1H, H7),
10.54 (br, NH benzimidazole, D2O exchangeable). IR (cmꢀ1): 3420.3
(NH benzimidazole), 3343.75 (NH hydrazide), 1670.62 (C]O),
1635.7 (C]N Schiff base), 1627.82 (C]N benzimidazole), 1524 and
1329.7 (NO2). MS: m/z 275 (Mþ, 12%). Anal. Calcd for C12H13N5O3: C,
52.36; H, 4.76; N, 25.44. Found: C, 52.41; H, 4.83; N, 25.35.
4.1.12. 2-Methyl-N-(4-methylbenzylidine)-6-nitro-1H-
benzimidazole-5-carbohydrazide (5g)
Green solid. Rf ¼ 0.46 (ethyl acetate/methanol, 4:1/4). Yield:
90%, m.p.140–142 ꢁC. 1H NMR (270 MHz, DMSO-d6)
d 2.1 (s, 3H, CH3
phenyl), 2.48 (s, 3H, CH3 benzimidazole), 6.95 (d, J ¼ 8.01 Hz, 2H,
H30 þ H50 phenyl), 7.31 (d, J ¼ 8.01 Hz, 2H, H20 þ H60 phenyl), 7.78
(s, 1H, NH hydrazide, D2O exchangeable), 8.07 (s, 1H, CH]N), 8.31
(s, 1H, H4 benzimidazole), 8.50 (s, 1H, H7), 9.03 (br, NH benz-
imidazole, D2O exchangeable). IR (cmꢀ1): 3486.75 (NH benzimid-
azole), 3375 (NH hydrazide), 1666.64 (C]O), 1635.62 (C]N Schiff
base), 1625 (C]N), 1525.42 and 1333.57 (NO2). MS: m/z 337 (Mþ,
15%). Anal. Calcd for C17H15N5O3: C, 60.53; H, 4.48; N, 20.76. Found:
C, 60.48; H, 4.40; N, 20.80.
4.1.8. N-((Furan-2-yl)methylene)-2-methyl-6-nitro-1
H-benzimidazole-5-carbohydrazide (5c)
Green solid. Rf ¼ 0.46 (ethyl acetate/methanol, 4:1/4). Yield:
85%, m.p. 285–287 ꢁC. 1H NMR (500 MHz, DMSO-d6)
d 2.45 (s, 3H,
CH3 benzimidazole), 6.3 (m, 1H, H40 furan), 6.71 (m, 1H, H30 furan),
7.2 (m,1H, H50 furan), 7.53 (s,1H, NH hydrazide, D2O exchangeable),
7.75 (s, 1H, CH]N), 8.03 (s, 1H, H4 benzimidazole), 8.61 (s, 1H, H7
benzimidazole), 9.34 (br, NH benzimidazole, D2O exchangeable). IR
(cmꢀ1): 3437.50 (NH benzimidazole), 3201.87 (NH hydrazide),
1664.07 (C]O), 1630.7 (C]N Schiff base), 1625.86 (C]N benz-
imidazole), 1518.25 and 1331.93 (NO2). MS: m/z 312 (Mþ ꢀ 1, 50%).
Anal. Calcd for C14H11N5O4: C, 53.68; H, 3.54; N, 22.36. Found: C,
53.60; H, 3.50; N, 22.31.
4.1.13. Preparation of the metal complexes (6–15)
General procedure. A solution of the metal ion in ethanol was
added to a solution of the corresponding ligand in ethanol to form
1:1 or 1:2 M:L complexes. The reaction mixture was stirred at 50 ꢁC,
for a time depending on the transition metal salt used. The resulting
precipitates were filtered off, washed with diethyl ether followed
by warm ethanol. The formed complexes were kept under dry
conditions.
4.1.9. 2-Methyl-N-(5-methylfuran-2-yl)methylene-6-nitro-1
H-benzimidazole-5-carbohydrazide (5d)
4.1.14. Cu–L1 complex [Cu(L1)Cl(H2O)]Cl$3H2O (6)
Green solid. Rf ¼ 0.44 (ethyl acetate/methanol, 4:1/4). Yield:
Green solid. Yield: 65%, m.p. 286–288 ꢁC, molar conductance Lm
93%, m.p. 159–161 ꢁC. 1H NMR (500 MHz, DMSO-d6)
d
2.09 (s, 3H,
(
U
ꢀ1 cm2 molꢀ1) ¼ 67. IR (cmꢀ1): 3580–3300 (OH water molecules,
CH3 furan), 2.49 (s, 3H, CH3 benzimidazole), 6.03 (d, J ¼ 8.1 Hz, 1H,
H40 furan), 6.44 (d, J ¼ 8.1 Hz, 1H, H30 furan), 7.50 (s, 1H, NH
hydrazide, D2O exchangeable), 7.65 (s, 1H, CH]N), 7.71 (s, 1H, H4
benzimidazole), 8.28 (s, 1H, H7 benzimidazole), 9.36 (br, NH
benzimidazole, D2O exchangeable). IR (cmꢀ1): 3421.87 (NH benz-
imidazole), 3328.12 (NH hydrazide), 1653.18 (C]O), 1633.98 (C]N
Schiff base), 1625.86 (C]N benzimidazole), 1521.45 and 1337.45
(NO2). MS: m/z 327 (Mþ, 10%). Anal. Calcd for C15H13N5O4: C, 55.05;
H, 4.00; N, 21.40. Found: C, 55.08; H, 4.12; N, 21.30.
NH benzimidazole), 3307.61–3155.80 (NH2, NH hydrazide), 2800–
2600 (intermolecular H-bonded NH), 1626.59 (C]N benzimid-
azole), 1608.00 (C]O coordinated). FT-IR (cmꢀ1): 691 (Cu–N), 691
(Cu–O). MS: m/z 396 (Mþ þ 1). Anal. Calcd for C9H18Cl2CuN4O5: C,
27.25; H, 4.57; Cl, 17.87; Cu, 16.02; N, 14.12. Found: C, 27.21; H, 4.41;
Cl, 17.91; Cu, 15.99; N, 14.09.
4.1.15. Ag–L1 complex [Ag(L1)NO3(H2O)] (7)
Pale brown solid. Yield: 75%, m.p. 234–235 ꢁC, molar conduc-
tance Lm
(U
ꢀ1 cm2 molꢀ1) ¼ 23. 1H NMR (500 MHz, DMSO-d6)
d 2.7
4.1.10. 2-Methyl-6-nitro-N-((pyridin-2-yl)methylene)-1
H-benzimidazole-5-carbohydrazide (5e)
(s, 3H, CH3), 3.66 (br, NH2, D2O exchangeable), 7.66 (d, J ¼ 8.5 Hz,
1H, H7), 7.79 (d, J ¼ 8.5 Hz, 1H, H6), 8.3 (s, 1H, H4), 9.63 (s, 1H, NH
hydrazide, D2O exchangeable), 10.3 (br, NH, D2O exchangeable). IR
(cmꢀ1): 3587–3386 (OH water molecules, NH benzimidazole),
3307.61–3143.12 (NH2, NH hydrazide), 2800–2600 (intermolecular
H-bonded NH), 1648.00 (C]O coordinated), 1625.36 (C]N). FT-IR
(cmꢀ1): 360 (Ag–N), 315 (Ag–O). MS: m/z 375 (Mþ ꢀ 2). Anal. Calcd
for C9H12AgN5O5: C, 28.59; H, 3.20; Ag, 28.53; N, 18.52. Found: C,
28.52; H, 3.17; Ag, 28.47; N, 18.49.
Yellow solid. Rf ¼ 0.43 (ethyl acetate/methanol, 4:1/4). Yield:
83%, m.p. 230–232 ꢁC. 1H NMR (500 MHz, DMSO-d6)
d 2.43 (s, 3H,
CH3 benzimidazole), 7.32 (s, 1H, NH hydrazide, D2O exchangeable),
7.60 (s, 1H, CH]N), 7.65 (m, 1H, H50 pyrimidine), 7.71 (s, 1H, H4
benzimidazole), 7.86 (m, 1H, H40 pyrimidine), 8.00 (m, 1H, H30
pyrimidine), 8.28 (s, 1H, H7 benzimidazole), 8.95 (m, 1H, H60
pyrimidine), 9.53 (br, NH benzimidazole, D2O exchangeable). IR
(cmꢀ1): 3418.33 (NH benzimidazole), 3223.38 (NH hydrazide),
1650.77 (C]O), 1632.98 (C]N Schiff base), 1625 (C]N), 1518.91
and 1384.32 (NO2). MS: m/z 324 (Mþ, 20%). Anal. Calcd for
C15H12N6O3: C, 55.55; H, 3.73; N, 25.91. Found: C, 55.60; H, 3.82; N,
25.79.
4.1.16. Ni–L1 complex [Ni(L1)Cl2(H2O)2]$H2O (8)
Dark brown solid. Yield: 82%, m.p. >300 ꢁC (decomposed
without melting), molar conductance Lm
(
U
ꢀ1 cm2 molꢀ1) ¼ 8.7. IR
(cmꢀ1): 3570–3350 (OH water molecules, NH benzimidazole),
3300–3164.12 (NH2, NH hydrazide), 2800–2600 (intermolecular H-
bonded NH), 1630.00 (C]O coordinated), 1625.37 (C]N). FT-IR
(cmꢀ1): 713 (Ni–N), 554 (Ni–O). Anal. Calcd for C9H16Cl2N4NiO4: C,
28.91; H, 4.31; Cl, 18.97; N, 14.99; Ni, 15.70. Found: C, 28.80; H, 4.33;
Cl, 18.85; N, 14.82; Ni, 15.79.
4.1.11. N-((1H-Indol-3-yl)methylene)-2-methyl-6-nitro-1
H-benzimidazole-5-carbohydrazide (5f)
Yellow solid. Rf ¼ 0.42 (ethyl acetate/methanol, 4:1/4). Yield:
87%, m.p. 268–270 ꢁC. 1H NMR (270 MHz, DMSO-d6)
d 2.59 (s, 3H,
CH3), 7.18–7.23 (m, 2H, H50 þ H60 indole), 7.28–7.31 (m, 2H,
H40 þ H70 indole), 7.66 (s, 1H, H4 benzimidazole), 7.67 (d,
J ¼ 2.14 Hz, 1H, H20 indole), 7.77 (s, 1H, NH hydrazide, D2O
exchangeable), 7.83 (d, J ¼ 2.14 Hz, 1H, CH]N), 8.12 (s, 1H, H7
4.1.17. Fe–L1 complex [Fe(L1)Cl3(H2O)]$3H2O (9)
Yellow solid. Yield: 71%, m.p. 218–219 ꢁC, molar conductance Lm
(U
ꢀ1 cm2 molꢀ1) ¼ 28. IR (cmꢀ1): 3592–3375 (OH water molecules,

S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
1507
NH benzimidazole), 3203.12–3155.80 (NH2, NH hydrazide), 2800–
2600 (intermolecular H-bonded NH), 1625 (C]N), 1613.12 (C]O
coordinated). FT-IR (cmꢀ1): 500 (Fe–N), 485 (Fe–O). MS: m/z 425
(Mþ þ 2). Anal. Calcd for C9H18Cl3FeN4O5: C, 25.47; H, 4.27; Cl,
25.06; Fe, 13.16; N, 13.20. Found: C, 25.43; H, 4.37; Cl, 25.00; Fe,
13.29; N, 13.11.
(intermolecular H-bonded NH), 1633.44 (C]O coordinated),
1623.44 (C]N benzimidazole), 1578.12 (C]N Schiff base). FT-IR
(cmꢀ1): 450 (Mn–N), 380 (Mn–O). Anal. Calcd for
C12H20Cl2MnN4O4: C, 35.14; H, 4.91; Cl, 17.29; Mn, 13.39; N, 13.66.
Found: C, 35.12; H, 4.81; Cl, 17.11; Mn, 13.44; N, 13.55.
4.2. Cytotoxicity assays
4.1.18. Mn–L1 complex [Mn(L1)2Cl(H2O)]Cl$3H2O (10)
Buff solid. Yield: 63%, m.p. >300 ꢁC (decomposed without
Cell culture cytotoxicity assays were carried out as described
melting), molar conductance Lm
(
U
ꢀ1 cm2 molꢀ1) ¼ 78. IR (cmꢀ1):
previously [26]. Briefly, aliquots of 100 mL cell suspension (1–
3 ꢃ 103 cells) were placed in microtiter plates in an atmosphere of
3587–3386 (OH water molecules, NH benzimidazole), 3287–
3146.25 (NH2, NH hydrazide), 2800–2600 (intermolecular H-
bonded NH), 1639.76 (C]O coordinated), 1625.00 (C]N). FT-IR
(cmꢀ1): 465 (Mn–N), 385 (Mn–O). Anal. Calcd for
C18H28Cl2MnN8O6: C, 37.38; H, 4.88; Cl, 12.26; Mn, 9.50; N, 19.38.
Found: C, 37.45; H, 4.92; Cl, 12.31; Mn, 9.40; N, 19.35.
5% CO2 at 37 ꢁC. After 24 h, 100
mL of culture media and 2 mL of the
compound in DMSO were added to each well in duplicate, and the
plates incubated for an additional 72 h at 37 ꢁC. There was no effect
on the growth of cells compared to that of cells in culture media
alone at this DMSO concentration. Compounds, along with mito-
mycin C as a positive control, were evaluated at final concentrations
4.1.19. Cu–L2 complex [Cu(L2)Cl2(H2O)2]$H2O (11)
ranging from 0.001 to 50
After the 72 h incubation, culture media were removed from
each well, and 200 L of fresh media and 20 L of AlamarBlue
mM.
Dark brown solid. Yield: 65%, m.p. 276–278 ꢁC (decomposed
without melting), molar conductance Lm
(
U
ꢀ1 cm2 molꢀ1) ¼ 15.8.
m
m
IR (cmꢀ1): 3589–3310 (OH water molecules, NH benzimidazole,
hydrazide), 2800–2600 (intermolecular H-bonded NH), 1635.86
(C]O coordinated), 1625.37 (C]N benzimidazole), 1578.00 (C]N
Schiff base). FT-IR (cmꢀ1): 711 (Cu–N), 563 (Cu–O). Anal. Calcd for
C12H20Cl2CuN4O4: C, 34.42; H, 4.81; Cl, 16.93; Cu, 15.17; N, 13.38.
Found: C, 34.41; H, 4.76; Cl, 16.91; Cu, 15.29; N, 13.29.
reagent were added, followed by additional 6 h incubation. Cell
viability was detected by fluorescent intensity using a Beckman
Coulter DTX880 plate reader with excitation at 530 nm and emis-
sion at 590 nm.
The fluorescence data obtained from the cytotoxicity studies
were used to calculate the percent growth according to the
following equation:
4.1.20. Ag–L2 complex [Ag(L2)2]NO3$H2O (12)
Pale brown solid. Yield: 60%, m.p. 219–220 ꢁC, molar conduc-
% Growth ¼ 100 ꢃ ðFtest ꢀ Ftime0Þ=ðFctrl ꢀ Ftime0
Þ
(1)
tance Lm
(
U
ꢀ1 cm2 molꢀ1) ¼ 65. 1H NMR (200 MHz, DMSO-d6)
where mean Ftime0 ¼ the averaged measured fluorescent intensities
of AlamarBlue reagent at the time just before the exposure of the
cells to the test substance, mean Ftest ¼ the averaged measured
fluorescent intensities of AlamarBlue reagent after 72 h exposure of
the cells to the test substance at a particular concentration, mean
Fctrl ¼ the averaged measured fluorescent intensities of AlamarBlue
reagent after 72 h exposure of the cells to the vehicle without the
test substance.
The IC50, the compound concentration for which the growth of
treated cells from time 0 was only 50% as much as the vehicle-
control was determined by non-linear regression fitting the data to
equation [2]:
d
2.10 (s, 6H, 2CH3 isopropylidine), 2.14 (s, 6H, 2CH3 isopropylidine),
2.73 (s, 6H, 2CH3 benzimidazole), 7.40 (br, NH hydrazide, D2O
exchangeable), 7.75 (d, J ¼ 8.5 Hz, 2H, H7 þ H70), 7.90 (d, J ¼ 8.5 Hz,
2H, H6 þ H60), 8.45 (s, 2H, H4 þ H40), 10.35 (br, NH benzimidazole,
D2O exchangeable). IR (cmꢀ1): 3580–3377 (OH water molecules,
NH benzimidazole), 3303.00 (NH hydrazide), 2800–2600 (inter-
molecular H-bonded NH), 1639.62 (C]O coordinated), 1627.39
(C]N benzimidazole), 1577.37 (C]N Schiff base). FT-IR (cmꢀ1):
356.86 (Ag–N), 318 (Ag–O). MS: m/z 649 (Mþ þ 2). Anal. Calcd for
C24H30AgN9O6: C, 44.46; H, 4.66; Ag, 16.64; N, 19.44. Found: C,
44.60; H, 4.57; Ag, 16.55; N, 19.40.
ꢀ
ꢁ
4.1.21. Ni–L2 complex [Ni(L2)2Cl2]$5H2O (13)
50ÞꢃnÞ
y ¼ min þ ðmax ꢀ minÞ= 1 þ 10ððxꢀlog IC
(2)
Green solid. Yield: 61%, m.p. >300 ꢁC (decomposed without
melting), molar conductance Lm
(U
ꢀ1 cm2 molꢀ1) ¼ 12.3. IR (cmꢀ1):
where min ¼ the minimum response plateau (0% growth),
max ¼ the maximum response plateau (100% growth), y ¼ %
growth at each test compound concentration, and n is a fitted
parameter (the Hill slope coefficient).
3570–3350 (OH water molecules, NH benzimidazole), 3305.25 (NH
hydrazide), 2800–2600 (intermolecular H-bonded NH), 1638.00
(C]O coordinated), 1625.00 (C]N benzimidazole), 1576.83 (C]N
Schiff base). FT-IR (cmꢀ1): 720.12 (Ni–N), 561.85 (Ni–O). Anal. Calcd
for C24H38Cl2N8NiO7: C, 42.38; H, 5.63; Cl, 10.42; N, 16.47; Ni, 8.63.
Found: C, 42.29; H, 5.57; Cl, 10.35; N, 16.41; Ni, 8.60.
Acknowledgement
We thank the US–Egypt Joint Science and Technology Board
Fund, administered through the USDA (BIO9-002-015).
4.1.22. Fe–L2 complex [Fe(L2)2Cl2]Cl$2H2O (14)
Brown solid. Yield: 67%, m.p. >300 ꢁC, molar conductance Lm
(U
ꢀ1 cm2 molꢀ1) ¼ 76. IR (cmꢀ1): 3590–3366 (OH water molecules,
NH benzimidazole), 3238.14 (NH hydrazide), 2800–2600 (inter-
molecular H-bonded NH), 1638.47 (C]O coordinated), 1625.00
(C]N benzimidazole), 1583.75 (C]N Schiff base). FT-IR (cmꢀ1):
520 (Fe–N), 472 (Fe–O). MS: m/z 639 (Mþ ꢀ H2O). Anal. Calcd for
C24H32Cl3FeN8O4: C, 43.76; H, 4.90; Cl, 16.15; Fe, 8.48; N, 17.01.
Found: C, 43.70; H, 4.93; Cl, 16.09; Fe, 8.40; N, 17.07.
Appendix Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.ejmech.2008.07.013.
References
[1] R.J. Sundberg, R.B. Martin, Chem. Rev. 74 (1974) 471–514.
[2] Q. Zhou, P. Yang, Inorg. Chim. Acta 359 (2006) 1200–1206.
[3] E. Lukevics, P. Arsenyan, I. Shestakova, I. Domracheva, A. Nesterova, O. Pudova,
Eur. J. Med. Chem. 36 (2001) 507–515.
4.1.23. Mn–L2 complex [Mn(L2)Cl2(H2O)2]$H2O (15)
Brown solid. Yield: 59%, m.p. >300 ꢁC (decomposed without
melting), molar conductance Lm
(U
ꢀ1 cm2 molꢀ1) ¼ 26. IR (cmꢀ1):
¨
3587–3386 (OH water molecules, NH benzimidazole), 2800–2600
[4] F. Gu¨mu¨s, O. Algu¨l, J. Inorg. Biochem. 68 (1997) 71–74.

1508
S.A. Galal et al. / European Journal of Medicinal Chemistry 44 (2009) 1500–1508
¨
¨
[5] F. Gu¨mu¨ s, O. Algu¨l, G. Eren, H. Erog˘lu, N. Diril, S. Gu¨r, A. Ozku¨l, Eur. J. Med.
Chem. 38 (2003) 473–480.
[16] K. Nakamoto, Infrared and Raman Spectra of Inorganic and Coordination
Compounds, fourth ed. John Wiley and Sons, 1986.
[17] D.J. Rabiger, M.M. Joullie, J. Org. Chem. 29 (1964) 476–482.
[18] R.C. Maurya, D.D. Mishra, Synth. React. Inorg. Met.-Org. Chem. 20 (8) (1990)
1013–1024.
[19] N. Bharti, M.T. Shailendra, G. Gonzalez, D. Garza, E. Cruz-Vega, J. Castro-Garza,
K.F. Saleem, M.R. Naqvi, Maurya, A. Azam, Bioorg. Med. Chem. Lett. 12 (2002)
869–871.
[20] M.M. Khalil, H.A. Mohamed, S.M. El-Medani, R.M. Ramadan, Spectrochim. Acta
59 (A) (2003) 1341–1347.
[21] A.A. Tak, F. Arjmand, S. Tabassum, Transition Met. Chem. 27 (2002) 741–747.
[22] J. Zhang, P. Braunstein, R. Welter, Inorg. Chem. 43 (2004) 4172–4177.
[23] F. Arjmand, B. Mohani, S. Ahmed, Eur. J. Med. Chem. 40 (2005) 1103–1110.
[24] N.S. Youssef, K.H. Hegab, Synth. React. Inorg. Met.-Org. Chem. 35 (5) (2005)
391–399.
_
¨
[6] F. Gu¨mu¨ s, F. Izgu¨, O. Algu¨l, FABAD J. Pharm. Sci. 21 (1996) 7–15.
[7] F. Gu¨mu¨s, I. Pamuk, T. Ozden, S. Yildis, N. Diril, S. Oksu¨zo¨glu¨, A. Gu¨ r, J. Ozku¨l,
Inorg. Biochem. 94 (2003) 255–262.
_
¨
¨
¨
¨
[8] M. Go¨kçe, S. Utku, S. Gu¨ r, A. Ozkul, F. Gu¨mu¨s, Eur. J. Med. Chem. 40 (2005)
135–141.
[9] A.M. Phillips, J. Chem. Soc. (1928) 2393–2399.
[10] N. Yamasaki, T. Imoto, Y. Murai, T. Hiramura, T. Oku, K. Sawada, PCT Int. Appl.
WO 97/24,334, JP Appl. 96/287,676. Chem. Abstr. 127 (1997) 135799d.
[11] I.M. Labouta, A.M.M. Hassan, O.M. Aboulwafa, O. Kader, Monatsh. Chem. 120
(6–7) (1989) 571–574.
[12] F.L. Mercha´n, J. Garı´n, E. Mele´ndez, D. Ortiz, T. Tejero, Synthesis (1987) 368.
[13] K.D.A. Domopoulou, M.A. Demertzis, A. Papageorgiou, J. Inorg. Biochem. 68
(1997) 147–155.
[14] S. Sharma, F. Athar, M.R. Maurya, A. Azam, Eur. J. Med. Chem. 40 (2005)
1414–1419.
[25] S.J. Berners-Price, R.J. Bowen, P. Galettis, P.C. Healy, M.J. McKeage, Coord.
Chem. Rev. 185–186 (1999) 823–836.
[15] N.S. Youssef, K.H. Hegab, A.E. Eid, Synth. React. Inorg. Met.-Org. Chem. 33 (9)
(2003) 1647–1666.
[26] D. Kumar, M.R. Jacob, M.B. Reynolds, S.M. Kerwin, Bioorg. Med. Chem. 10
(2002) 3997–4004.