Bis-aroylhydrazone based on 2,2′-bis substituted diphenylamine for synthesis of new binuclear organotin (IV) complexes: Spectroscopic characterization, crystal structures, in vitro DNA-binding, plasmid DNA cleavage, PCR and cytotoxicity against MCF7 cell

Article abstract of DOI:10.1002/aoc.5137

New dinuclear organotin (IV) complexes, Me₄Sn₂L, Ph₄Sn₂L and Bu₄Sn₂L, have been synthesized from reaction of R₂SnCl₂ (R?=?Me, Ph and Bu) with a 2,2′-bis-substituted di

Full text of DOI:10.1002/aoc.5137

Received: 10 January 2019
DOI: 10.1002/aoc.5137
Revised: 1 July 2019
Accepted: 2 July 2019
F U L L P A P E R
Bis‐aroylhydrazone based on 2,2′‐bis substituted
diphenylamine for synthesis of new binuclear organotin
(IV) complexes: Spectroscopic characterization, crystal
structures, in vitro DNA‐binding, plasmid DNA cleavage,
PCR and cytotoxicity against MCF7 cell line
Maryam Yousefi1 | Tahereh Sedaghat1
| Jim Simpson2 | Mohammad Shafiei3
1 Department of Chemistry, Faculty of
New dinuclear organotin (IV) complexes, Me4Sn2L, Ph4Sn2L and Bu4Sn2L,
have been synthesized from reaction of R2SnCl2 (R = Me, Ph and Bu) with a
2,2′‐bis‐substituted diphenylamine arοylidene hydrazone, H4L. The synthe-
Science, Shahid Chamran University of
Ahvaz, Ahvaz, Iran
2 Department of Chemistry, University of
Otago, PO Box 56, Dunedin 9054, New
Zealand
1
sized compounds were investigated by elemental analysis and infrared, H‐
NMR and 119Sn‐NMR spectroscopy. The structures of H4L, Me4Sn2L and
Bu4Sn2L were also confirmed by X‐ray crystallography. H4L molecules adopt
(E)‐configuration and keto‐tautomeric form in the solid state. In all complexes,
the bis‐hydrazone acts as a tetra‐anionic ligand with two contiguous ONO
tridentate domains that coordinate the two R2Sn moieties in the enolate form.
The coordination environments of both tin centers are five‐coordinate. DNA‐
binding studies were performed by UV–Vis spectroscopy, and the results indi-
cate that the synthesized compounds significantly interact with calf thymus‐
DNA in the intercalative mode. The results of polymerase chain reaction assay
show that all the compounds affect on amplification of DNA, and complexes
are more effective than ligands. The in vitro cytotoxicity against the human
breast cancer line (MCF7) was determined using the MTT assay, and H4L
and the dibutytin complex showed higher activity.
3 Department of Biology, Faculty of
Science, Shahid Chamran University of
Ahvaz, Ahvaz, Iran
Correspondence
Department of Chemistry, Faculty of
Science, Shahid Chamran University of
Ahvaz. Ahvaz, Iran.
Email: sedaghat@scu.ac.ir
Funding information
Shahid Chamran University of Ahvaz,
Grant/Award Number: 1396
KEYWORDS
119Sn NMR, crystal structure, DNA, hydrazone, organotin (IV)
1 | INTRODUCTION
synthesized and their anti‐cancer properties tested. These
compounds could be as effective as, or even better than,
Recently, a great deal of research has been done in the
field of inorganic and organometallic chemistry targeting
the preparation of metal‐based drugs. The purpose of
these investigations is to find more effective drugs with
fewer side‐effects by changing ligands or metals in the
complex molecules.[1] Among these target systems,
organotin (IV) complexes have been of great interest,
and so far a large number of these compounds have been
traditional heavy metal anticancer drugs such as the
platin complexes.[2–10] A smart choice of the ligand coor-
dinated to the organotin (IV) moiety can increase the
activity of the complex or decrease its harmful side‐effects
or cross‐resistance.[11,12] In the meantime, hydrazones
have received much attention due to their potential bio-
logical applications, including antitumor activity, antibac-
terial properties and iron‐chelating ability. In
Appl Organometal Chem. 2019;e5137.
wileyonlinelibrary.com/journal/aoc
© 2019 John Wiley & Sons, Ltd.
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https://doi.org/10.1002/aoc.5137

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YOUSEFI ET AL.
coordination chemistry, hydrazones that include a range
of potential donor groups are recognized as a very impor-
tant class in multi‐dentate ligands. They also have the
ability to form mono‐, di‐ or multi‐nuclear complexes
with both main group and transition metal centers.
Acyl/arοyl‐hydrazones with the ‐C=NNHC(O)‐ func-
tional group are a particular category of these type of
hydrazone compounds, and interest in their use as multi-
functional ligands has increased considerably.[13,14]
Among these derivatives, the chemistry of bis
acyl/arοyl‐hydrazone compounds is seen to be of particu-
lar interest for the following reasons: (a) there is the pos-
sibility of creating supermolecular structures and various
types of coordination polymers using these ligands; (b)
coordination of a metal ion into such a unit may cause
changes in the characteristics of the other units; (c) due
to the different degree of deprotonation available for
these multi‐proton ligands, complexes with various
charges can be synthesized.[15,16] Bis‐acyl/arοyl‐
hydrazones can be prepared from the reaction of a
dihydrazide and an aldehyde or ketone in a 1:2 ratio, or
from a similar reaction of a dialdehyde or diketone with
a hydrazide. It is visualized that the flexibility of distance
introduced in the linkers of these bifunctional
hydrazones may result in interesting structural or reactiv-
ity features in the dihydrazone and its metal complexes.
Recently, the synthesis and structural study of some
binuclear organotin (IV) complexes of bis‐acyl/arοyl‐
hydrazones have been reported,[17–20] and we have also
published in this field.[21–26] In the present investigation,
we have used diphenylamine derived from anthranilic
acid as a linker for the synthesis of a bis‐hydrazone and
its dinuclear Sn (IV) complexes. N‐Aryl anthranilic acid
derivatives are valuable compounds with pain‐killing,
antipyretic and anti‐inflammatory properties.[27] Further-
more, recent researches have demonstrated that deriva-
tives of diphenylamine can act as kinase inhibitors.[28,29]
Kinases are well‐known regulators of many cellular path-
ways, so a significant proportion of human proteins can
be modified by kinase activity. Therefore, mutation or
excessive expression of kinase proteins plays a fundamen-
tal role in diverse cellular activities such as growth and
proliferation. As a result, a great deal of attention has
been paid to compounds that act as kinase inhibitors,
and consequentially can be effective as anti‐proliferative
and/or cytotoxic agents.[30–32] Quinazoline‐based mole-
cules are another class of these compounds with inhibi-
tory properties against tumor cell proliferation.[33–35]
Owing to the popularity of hydrazone anticancer activity,
a
series of hydrazones containing diphenylamines
substituted by 2,2′‐bis arοylhydrazide have been synthe-
sized to imitate the pyrimidine ring in quinazoline. Such
a substitution pattern could target different ATP‐binding
sites. The scaffolds formed by intramolecular hydrogen
bonds between the NH group of the diphenylamine and
the two hydrazide oxygen atoms effectively mimic the
quinazoline kinase template by forming a pseudo six‐
member ring.[28] Based on this knowledge and in a con-
tinuation of our work on binuclear organotin complexes
of bis‐hydrazones, in this study a 2,2′‐bis‐substituted
diphenylamine arοylidene hydrazine (Figure 1) was syn-
thesized. A halogenated derivative was chosen with the
aim of improving the activity and lipophilicity of the
product.[27] This bis‐hydrazone was then used as a
binucleating ligand for the synthesis of new organotin
(IV) complexes.
2 | EXPERIMENTAL
2.1 | Materials and methods
The materials and solvents in this research work were
obtained from commercial suppliers and used as received
without further purification. Infrared (IR) spectra of syn-
thesized compounds were recorded on a FT BOMEM
MB102 spectrophotometer as KBr discs. The 1H‐ and
119Sn‐NMR spectra were recorded with a Bruker Avance
Ultrashield spectrometer using TMS and SnMe4, respec-
tively, as internal references.
FIGURE 1 Synthetic pathway to the
ligand and their complexes

YOUSEFI ET AL.
3 of 15
2.2 | Synthesis of 2‐(2‐
carboxyphenylamino) benzoic acid (1)
2.4 | Synthesis of 2‐[2‐(hydrazinecarbonyl)
phenylamino] benzohydrazide (3)
This compound was synthesized by a similar procedure to
that reported for the 2,4′‐substituted derivative with some
modifications.[36] Potassium carbonate (15 g, 180 mmol)
was added to 2‐chlorobenzoic acid (5.16 g, 33 mmol) in
dimethylformamide (40 ml) and stirred for 15 min.
Separately, a solution of anthranilic acid (4.93 g,
36 mmol) and potassium carbonate (15 g) in DMF
(20 ml) was stirred for 15 min, and then activated copper
powder (0.6 g) was added. This mixture was allowed to
stir about 15 min (the copper powder was activated by
washing with a 1:l mixture of acetone and concentrated
HCl and then with aqueous acetone, and dried in a vac-
uum oven). The two mixtures with an additional amount
of dimethylformamide (30 ml) were mixed together and
refluxed for 20 hr at 140–145°C and under a nitrogen
gas atmosphere.
Hydrazine hydrate (64 ml) was added to compound 2
(6.7 g, 23.6 mmol) and the solution was refluxed for
15 hr. During this time, the color changed from light
green to white, and then the resulting white solid was fil-
tered and dried. Yield: 6.2 g (95%); m.p. 273–275°C. FT‐IR
(KBr, cm−1): ν(N‐H), 3360, 3278, 3172; ν(C=O), 1645. 1H‐
NMR (300 MHz, DMSO‐d6): δ 10.22 (s, 1H, NH), 9.64 (s,
2H, NH), 7.48–6.84 (m, 8H, Ar‐H), 4.44 (s, 4H, NH2).
2.5 | Synthesis of H4L (4)
A solution of 5‐bromo‐2‐hydroxybenzaldehyde (0.402 g,
2.5 mmol) and the dihydrazide (3) (0.317 g, 1.2 mmol) in
MeOH (13 ml) was refluxed for 5 hr. The solid product
was filtered, washed with MeOH and dried in an oven at
50°C. Colorless block‐like crystals suitable for X‐ray crys-
tallography were obtained from slow evaporation of an
ethanol/chloroform (3:1) solution. Yield: 0.37 g (48%); m.
p. 224–226°C. FT‐IR (KBr, cm−1): ν(O‐H), 3436; ν(N‐H),
During this time the color of the mixture changed from
brown to red then to pink. Next the mixture was cooled to
60°C, added to 200 ml of ice/water and stirred for 15 min.
This solution was decolorized using activated carbon (3 g)
at 100°C with stirring for 10 min. The mixture was filtered
and then HCl (6 M) was added to the solution in an ice‐
bath until the formation of precipitation was completed.
The yellow solid product was filtered, washed with water
and dried at 50°C in an oven. Yield: 6.62 g (78%); m.p.
293–295°C; Fourier transform (FT)‐IR (KBr, cm−1): ν(N‐
1
3219; ν(C=O), 1646; ν(C=N), 1609. H‐NMR (300 MHz,
DMSO‐d6): δ 12.18 (s, 2H, OH), 11.27 (s, 2H, NH), 10.22
(s, 1H, NH), 8.53 (s, 2H, CH=N), 7.77–6.84 (m, 14H, Ar‐H).
2.6 | Synthesis of [Me4Sn2L] (5)
1
H), 3330 sh; ν(O‐H), 3000–2800 br; ν(C=O), 1667. H‐
To a warm stirred solution of H4L (0.162 g, 0.20 mmol) in
methanol (13 ml), triethylamine (0.14 ml, 1 mmol) was
added. This suspension was stirred for 15 min, and then
Me2SnCl2 (0.109 g, 0.5 mmol) in methanol (5 ml) was
added. The color of the mixture changed from yellow to
red. This mixture was refluxed for 3 hr, and then the
orange precipitate was collected, washed with methanol
(5 ml) and dried at room temperature. Red plate‐like crys-
tals suitable for X‐ray crystallography were obtained from
slow evaporation of an ethanol/chloroform (3:1) solution.
Yield: 0.21 g (93%); m.p. 253–256°C. Anal. calcd for
C32H29Br2N5O4Sn2: C, 40.67; H, 3.09; N, 7.41%. Found:
C, 40.72; H, 2.64; N, 7.71%. FT‐IR (KBr, cm−1): ν(N‐H),
NMR (300 MHz, DMSO‐d6): δ 13.01 (s, 2H, OH), 10.81(s,
1H, NH), 7.91–6.91 (m, 8H, Ar‐H).
2.3 | Synthesis of methyl‐2‐[2‐
(methoxycarbonyl)phenylamino] benzoate
(2)
This compound was obtained by esterification of 1 by a
method similar to that reported for the chloro‐analog
with some modifications.[27] A solution of 1 (6.20 g,
24.12 mmol) in methanol (125 ml) was stirred in an ice‐
bath, and thionyl chloride (6.1 ml) was added dropwise
to it. The mixture was stirred at room temperature for
20 min, and then refluxed for 12 hr. After this time, the
dark solution was reduced to less than half the initial vol-
ume and left to cool. Pale green cotton‐like crystals
formed were collected and dried in an oven at 50°C.
Yield: 6.4 g (98%); m.p. 90–92°C; FT‐IR (KBr, cm−1):
ν(N‐H), 3309 sh; ν(C‐Haliphatic), 2985–2940; ν(C=O),
1700. 1H‐NMR (300 MHz, DMSO‐d6): δ 10.72(s, 1H,
NH), 7.92–6.95 (m, 8H, Ar‐H), 3.85 (s, 6H, CH3).
1
3223; ν(C=N), 1605; ν (Sn‐O), 564; ν (Sn‐N), 421. H‐
NMR (300 MHz, DMSO‐d6): δ 11.06 (s, 2H, NH), 8.58 [s,
2H, CH=N, 3J(119Sn‐1H) = 31.02 Hz], 7.94–6.56 (m,
14H, Ar‐H), 0.69 [s, 12H, Me2Sn, 2J(119Sn‐
1H) = 90.6 Hz]. 119Sn‐NMR (111 MHz, DMSO): δ −248.4.
2.7 | Synthesis of [Ph4Sn2L] (6)
Complex 6 was synthesized as described for 5 from H4L
(0.162 g, 0.25 mmol), triethylamine (0.13 ml, 1 mmol)

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YOUSEFI ET AL.
and Ph2SnCl2 (0.172 g, 0.0.5 mmol) in ethanol (15 ml).
Yield: 0.10 g (40%); m.p. 234–236°C. Anal. calcd for
C52H37Br2N5O4Sn2: C, 52.35; H, 3.13; N, 5.87%. Found:
C, 52.56; H, 3.00; N, 6.15%. FT‐IR (KBr, cm−1): ν(N‐H),
peaks remained in the difference Fourier map that
appeared to represent four water molecules. Three of
these refined sensibly and it was possible to locate their
H atoms in a difference Fourier map, and their coordi-
nates were refined with Ueq = 1.5 Ueq(O). The fourth sol-
vate atom did not refine reasonably with high and
increasing thermal parameters, suggesting significant dis-
order. The contribution to the scattering from this solvent
was therefore removed with the SQUEEZE routine[41]
within PLATON,[42] producing a solvent accessible void
of 818 Å3.
1
3260; ν(C=N), 1606; ν (Sn‐O), 576; ν (Sn‐N), 446. H‐
NMR (300 MHz, DMSO‐d6): δ 11.27 (s, 2H, NH), 8.42 [s,
3
2H, CH=N, J(119Sn‐1H) = 45.0 Hz], 8.19–6.79 (m, 14H,
Ar‐H). 119Sn‐NMR (111 MHz, DMSO): δ −434.2. Our
attempts for preparation of crystals suitable for X‐ray
crystallography were unsuccessful.
The SQUEEZE routine was also used for the refine-
ment of 5, which displayed what appeared to be a
severely disordered ethanol solvent molecule for which
again no sensible disorder model could be derived. This
produced a solvent accessible volume of 114 Å3 and fur-
ther refinement resulted in a significant reduction in the
final residuals.
2.8 | Synthesis of [Bu4Sn2L](7)
Complex 7 was synthesized as described for 5 from H4L
(0.162 g, 0.25 mmol), triethylamine (1 mmol) and
Bu2SnCl2 (0.152 g g, 0.5 mmol) in ethanol after 4 hr
reflux. Orange plate‐like crystals suitable for X‐ray crys-
tallography were obtained after a few days by slow evap-
oration of an ethanol solution. Yield: 0.13 g (48%); m.p.
74–76°C. Anal. calcd for C44H53Br2N5O4Sn2: C, 47.48; H,
4.80; N, 6.29%. Found: C, 47.51; H, 4.70; N, 6.26%. FT‐
IR (KBr, cm−1): ν(N‐H), 3229; ν(C‐H), 2955–2853;
ν(C=N), 1604; ν (Sn‐O), 566; ν (Sn‐N), 448. 1H‐NMR
(300 MHz, DMSO‐d6): δ 10.80 (s, 2H, NH), 8.67 [s, 2H,
The asymmetric unit of 7 consisted of two unique mol-
ecules 7.1 and 7.2. High and increasing displacement
parameters for the C atoms of the C42 and C43 methy-
lene and the adjacent C44 methyl groups of one butyl
chain in molecule 7.2 suggested positional disorder. Var-
ious disorder models were attempted, without success,
suggesting that the disorder spanned three or even more
locations. Finally the offending butyl chain was refined
with all atoms at single occupancy suing DFIX and ISOR
restraints. This procedure produced the best residuals of
the variety of models attempted, but still resulted in B
alerts in CheckCIF relating to high residual electron den-
sity and Hirshfeld bond distance alerts in the vicinity of
this butyl chain, suggesting further unresolvable disorder.
Less surprisingly, B alerts were also raised over high
residual density close to all four Sn (IV) atoms. A final
B alert, noting a close Br21 … O11 contact, simply signals
an unremarkable intermolecular Br … O halogen bond
that is mentioned in the description of the crystal packing
for this molecule.
3
CH=N, J(119Sn‐1H) = 27.1 Hz], 7.88–6.57 (m, 14H, Ar‐
H), 1.45–1.32 (m, 16H, CH2CH2), 1.16–1.10 (m, 8H,
3
CH2), 0.63 (t, 12H, CH3, JHH = 7.2 Hz). 119Sn‐NMR
(111 MHz, DMSO): δ −267.5.
2.9 | X‐ray structure determination
Colorless block‐like crystals of 4, a red plate of 5 and an
orange plate of 7 were used for data collection. X‐ray
measurements for all three structures were carried out
on an Agilent Duo diffractometer using CuKα radiation
(λ = 1.54184 Å) with data collection, reduction and
absorption corrections controlled using CrysAlisPro,[37]
and data were collected at 100(2) K. Data were reduced
and multi‐scan absorption corrections were applied using
CrysAlisPro.[37] All structures were solved by direct
methods using SHELXT,[38] and refined using full‐matrix
least‐squares procedures using SHELXL‐2018/1[39] and
TITAN2000.[40] All non‐hydrogen atoms were refined
anisotropically, and all hydrogen atoms bound to carbon
were placed in the calculated positions, and their thermal
parameters were refined isotropically with Ueq = 1.2–1.5
Ueq(C). The N‐H hydrogen atoms were located in differ-
ence Fourier maps, and their coordinates were refined
with Ueq = 1.2 Ueq(N).
Molecular plots and packing diagrams were drawn
using Mercury.[43] Additional metrical data were calcu-
lated using PLATON, and tabular material was produced
using WINGX.[44] Details of the X‐ray measurements and
crystal data for both complexes are given in Table 1.
2.10 | DNA‐binding
The binding affinity between calf thymus‐DNA (CT‐
DNA) and synthesized compounds was investigated by
UV–Vis spectroscopic titration in Tris–HCl/NaCl buffer
[5 mM Tris (hydroxymethyl)‐aminomethane; 50 mM
NaCl, pH 7.2] at room temperature. The solutions of
tested compounds were prepared in DMSO, diluted with
In addition to the three unique molecules of 4 in the
asymmetric unit, once all of the non‐hydrogen atoms
had been located and refined, several additional high

YOUSEFI ET AL.
5 of 15
TABLE 1 Crystallographic and structure refinement data for 4, 5 and 7
H4L·H2O (4)
[Me4Sn2L] (5)
[Bu4Sn2L] (7)
Empirical formula
Molecular weight
Temperature
C28H23Br2N5O5
C32 H29Br2N5O4Sn2
944.8
C44H53Br2N5O4Sn2
1113.11
669.33
100 K
100 K
100 (2) K
1.54184
Wavelength
1.54184
1.54184
Crystal system
Space group
monoclinic
Triclinic
Triclinic
P ‐1
I 2/a
P ‐1
a (Å)
29.3867(10)
11.5950(5)
12.3056(6)
14.1188(5)
86.436(3)
69.294(4)
64.069(5)
1684.41(15)
2
13.2746(3)
13.9614(4)
26.1209(5)
94.928(2)
93.291(2)
112.212(2)
4443.97(19)
4
b (Å)
15.3979(5)
c (Å)
37.6286(11)
α
90
β
104.000(3)
γ
90
Volume
16 520.9(9)
Z
24
Density (calculated)
Absorption coefficient
F (000)
1.615
1.863
1.664
4.143
14.934
11.420
8064
916
2216
Crystal size
0.29 × 0.11 × 0.05
3.441–74.482
0.20 × 0.09 × 0.05
3.367–73.575
31 651
0.32 × 0.18 × 0.07
3.413–75.003
32 981
Theta range for data collection°
Reflections collected
Independent reflections
Refinement method
Data/restraints/parameters
Goodness‐of‐fit on F 2
Final R indices [I > 2sigma(I)]
R indices (all data)
Largest diff. peak and hole (e/Å3)
32 950
16 264 [R (int) = 0.0490]
Full‐matrix least‐squares on F 2
16 264/12/1144
0.998
6714 [R (int) = 0.0784]
14 536 [R (int) = 0.0516]
6714/0/413
17 683/0/1060
1.044
1.025
R1 = 0.0475, wR2 = 0.1243
R1 = 0.0540, wR2 = 0.1315
1.661, −0.926
R1 = 0.0460, wR2 = 0.1160
R1 = 0.0574, wR2 = 0.1262
1.515, −1.036
R1 = 0.0561, wR2 = 0.1441
R1 = 0.0682, wR2 = 0.1544
2.334, −1.759
buffer, and an equal volume was added to DNA solutions
with various amounts of DNA in buffer (0, 0.5, 2, 5, 6 and
8 μg). The compound–DNA solutions were incubated at
room temperature for 5 min before the absorption spectra
were recorded.
2.12 | Polymerase chain reaction
investigation
Polymerase chain reaction (PCR) assays were monitored
to evaluate the interference of the compound in human
DNA amplification. In this method, interaction between
different concentration of compounds and human geno-
mic DNA was considered. The human genome was
extracted from blood samples and genome was used as
template in the PCR reaction. In this assay, genomic
DNA was treated with increasing concentrations of the
ligand and complexes (5, 25 and 100 μM). After incuba-
tion for 1 hr at room temperature, the PCR reaction mix-
ture was added and amplification of DNA was performed
on a thermal cycler. Ethidium bromide (5 and 25 μM) and
untreated genomic DNA were used as positive and nega-
tive controls, respectively. Then the PCR product was
2.11 | DNA cleavage
The DNA‐cleavage studies of compounds were performed
by agarose gel electrophoresis experiments, which were
done on supercoiled plasmid DNA (pET28b of
Escherichia coli; 0.1 μg/μl) in Tris–HCl buffer (10 μM,
pH 7.2) treated with the compounds at 37°C. The samples
were incubated for 3 hr, and then electrophoresed on 1%
agarose gel in TBE (Tris–boric acid–EDTA, pH 8.0) for
1 hr at 60 V.

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YOUSEFI ET AL.
electrophoresed on agarose gels at 60 V at 60 min and the
bands visualized with UV light.
hydroxybenzaldehyde afforded the bis‐hydrazone ligand,
2,2′‐azanediylbis(N′‐((E)‐5‐bromo‐2‐hydroxybenzylidene)
benzohydrazide) (4). The presence of the two ketoamide
units (NH‐C=O) in this structure allows the possibility
that keto‐enol tautomerism could occur in this structure
(Figure 2). The X‐ray structures, vide infra, reveal that,
while the H4L molecule adopts the keto form in the solid
state, complex formation is accompanied by adoption of
the enol tautomeric form, accompanied by deprotonation
of the resulting four OH substituents. Herein, we report
the molecular and crystal structure and nature of the
intramolecular hydrogen bonds for H4L.
Three new binuclear organotin (IV) complexes were
synthesized by reaction of H4L with R2SnCl2 (R = Me,
Ph and n‐Bu) in the presence of triethylamine in 1:2:4
ratio. The new complexes were characterized by elemen-
tal analysis and spectroscopic methods. The structures of
the methyl and butyltin (IV) complexes have been con-
firmed by X‐ray crystallography. The biological activities
of ligand and its three Sn (IV) complexes have also been
evaluated.
2.13 | MTT cytotoxicity assay
The cytotoxic activity of synthesized ligand and com-
plexes on the breast cancer cell line (MCF7) was exam-
ined by MTT assay [MTT: 3‐(4,5‐dimethylthiazol‐2‐yl)‐
2,5‐diphenyltetrazolium bromide]. Five‐thousand cells
were seeded in the wells of a 96‐well plate. Then, 100 μl
of various concentrations of synthesized compounds (25,
50, 75, 100 μM) in DMSO were added to the cells in trip-
licate. After 48 hr of incubation, the contents of the plate
were pipetted out carefully, cells were washed with
phosphate‐buffered saline (PBS) and fresh relevant
medium containing 20 μl of MTT solution in PBS
(0.5 mg/ml) was added to the test wells. The plate was
further incubated in a CO2 incubator at 37°C for 4 hr,
and after that time MTT assay was carried out. The assay
is based on reducing the tetrazolium salt MTT to insolu-
ble purple formazan by metabolically active cells that
are quantifiable by spectrophotometry. For this purpose,
the formed formazan crystals were dissolved in 120 μl of
DMSO for each well. The cell viability was determined
by measuring the absorbance of each well at 570 nm
and a reference wavelength of 630 nm using an ELISA
spectrophotometer.
3.1 | X‐ray structures of H4L·H2O,
[Me4Sn2L] and [Bu4Sn2L]
The asymmetric unit of the H4L compound comprises
three unique H4L molecules, identified in their number-
ing scheme by leading 1, 2 and 3 characters, and three
solvent water molecules. Figure 2 shows separate views
of the independent molecules 1–3, their atom numbering
and the intramolecular N‐H … O and O‐H … N hydrogen
bonds that they form. Table 2 gives the selected bond dis-
tances and angles for molecule 4.1. As mentioned previ-
ously, all three of the unique molecules in the solid
state structures adopt the keto‐tautomeric forms with
C18 = O12 and C121 = O13 carbonyl groups for molecule
4.1 and equivalent carbonyl substituents for the other two
molecules. All three molecules adopt an (E)‐configuration
about the ‐C17 = N11‐ and equivalent double bonds.
3 | RESULTS AND DISCUSSION
The synthetic pathway to the ligand (H4L, 4) is shown in
Figure 1. 2,2′‐Diphenylamine dicarboxylic acid (1) was
prepared from the reaction of 2‐chlorobenzoic acid with
anthranilic acid. Esterification of the product yielded
2‐[2‐(methoxycarbonyl)phenylamino]
methyl ester (2), which was then subjected to
hydrazinolysis to give 2‐[2‐(hydrazinecarbonyl)
phenylamino] benzoic acid hydrazide (3). Condensation
of the dihydrazide with 5‐bromo‐2‐
benzoic
acid
3
FIGURE 2 The three unique molecules of 4 showing the atom numbering and with ellipsoids drawn at the 50% probability level.
Intramolecular hydrogen bonds are shown as dotted lines. The three solvent water molecules found in this structure are not included

YOUSEFI ET AL.
7 of 15
TABLE 2 Bond lengths (Å) and angles (°) for complex ligand and complexes
H4L·H2O
O12‐C18
1.238(4)
N11‐C17
1.289(4)
C18‐N12‐N11
117.7(3)
C18‐N12
1.357(4)
1.377(4)
C14‐O11
1.353(4)
121.3(3)
N11‐C17‐C13
119.1(3)
130.6(3)
N12‐N11
N12‐C18‐O12
C114‐N13‐C115
Me4Sn2L
Sn1–O1
2.099(4)
2.159(4)
2.169(5)
97.4(2)
Sn1 –C15
2.114(6)
2.109(6)
1.294(7)
83.83(17)
106.8(2)
126.2(2)
72.32(17)
123.2(5)
111.5(5)
C8‐N2
1.326(8)
Sn1–O2
Sn1–C16
N2‐N1
1.389(6)
1.305(8)
Sn1–N1
O2‐C8
N1‐C7
O1–Sn1–C16
O1–Sn1–C15
C16–Sn1–C15
O1–Sn1–O2
C16–Sn1–O2
C15–Sn1–O2
Bu4Sn2L
O1–Sn1–N1
C16–Sn1–N1
C15–Sn1–N1
O2–Sn1–N1
O2–C8–N2
C8–N2–N1
N1–C7–C1
Sn1–O1–C2
Sn1–N1–C7
Sn1–N1–N2
Sn1–O2–C8
C14‐N3‐C14’
126.7(5)
94.2(2)
132.7(4)
128.4(4)
116.6(4)
114.4(4)
127.3(5)
126.6(2)
154.98(17)
96.7(2)
93.9(2)
Sn11‐O11
2.095(5)
2.158(5)
2.161(5)
92.9(2)
Sn11‐C133
2.121(6)
2.125(7)
1.308(8)
82.57(19)
120.2(2)
108.6(2)
72.52(19)
124.5(6)
110.1(5)
C18‐N12
1.318(9)
Sn11‐O12
Sn11‐C129
N12‐N11
1.407(8)
1.315(8)
Sn1 ‐N11
O12‐C18
N11‐C17
O11‐Sn11‐C133
O11‐Sn11‐C129
C133‐Sn11‐C129
O11‐Sn11‐ O12
C133‐Sn11‐O12
C129‐Sn11‐O12
O11‐Sn11‐N11
C133‐Sn11‐N11
C129‐Sn11‐N11
O12‐Sn11‐N11
O12–C18–N12
C18–N12–N11
N11–C17–C13
Sn11–O11–C14
Sn11–N11–C17
Sn11–N11–N12
Sn11–O12–C18
C115‐N13‐C114
125.9(6)
96.5(2)
133.7(4)
128.7(4)
117.8(4)
114.4(4)
128.0(6)
131.1(3)
154.16(18)
93.6(2)
97.9(2)
In molecules 4.1 and 4.3, the carbonyl groups form
two intramolecular N‐H … O hydrogen bonds with the
N‐H group of the central diphenylamine segment of the
molecule, each enclosing S6 ring motifs.[45] In contrast,
molecule 4.2 forms only a single intramolecular N23‐
H23 … O22 hydrogen bond that generates a similar ring
motif. Stronger O‐H … N intramolecular contact forms
for all three molecules between the OH substituents on
the hydroxybromobenzene rings and the adjacent imine
N atom, again enclosing S6 rings. Molecules 4.1 and 4.3
adopt similar, reasonably symmetrical, conformations,
and overlay[43] with an rms deviation of 0.48 Å. The two
benzene rings of the central diphenylamine ring systems
are inclined to one another at similar dihedral angles,
29.82(7)° for molecule 1 and 32.17(7)° for molecule 3. In
sharp contrast, the corresponding angle for molecule 2
is 58.87(9)°, destroying the pseudo mirror symmetry
found in the other two molecules and obviating the for-
mation of a pair of N‐H … O contacts by the N23 atom.
The markedly different conformation of 2 is also reflected
in the results of the overlay calculations with deviations
of 2.79 Å for the overlay of molecules 1 and 2, and
2.82 Å for 2 and 3, respectively. Within each individual
molecule, comparable bond distances do not differ signif-
icantly. The same can be said when comparing the three
unique molecules, except for those parameters associated
with the diphenylamine unit in molecule 2, which clearly
reflects the much greater skew of the unit's benzene rings
in that molecule. Bond distances and angles in the diphe-
nylamine segment of the molecule are similar to those in
related compounds.[46] However, no instances of struc-
tures of diphenylamine derivatives with O=CNH substit-
uents at the 2‐positions of the benzene rings are found in
the Cambridge Structural Database, Version 5.39, Nov.
2017 with 3 updates,[47] highlighting the exclusive nature
of the ligand reported here. The database also reveals
structures
of
four
(E)‐N′‐(5‐bromo‐2‐
hydroxybenzylidene)benzohydrazides, including the
archetypal hydrazide itself,[48] with the three others[49–
51] having various substituents on the benzene ring bound
to the amide unit. A related tin (IV) complex, dimethyl‐(N
′‐(5‐bromo‐2‐hydroxybenzylidene)benzoylhydrazone) tin
(IV), was also found.[52] In the crystal of 4, an extensive
series of classical O‐H … O, O‐H … N and N‐H … N

8 of 15
YOUSEFI ET AL.
hydrogen bonds (Table S1) combine with non‐classical C‐
H … Br and C‐H … O contacts to link the three unique
H4L molecules and the three water molecules into an
extensive three‐dimensional network (Figure 3).
The structures of complexes 5 and 7 were also deter-
mined from X‐ray diffraction data, and are sufficiently
similar to be discussed together. Both complexes crystal-
lize in the triclinic space group P‐1 with 2 and 4 mole-
cules in a unit cell of 5 and 7, respectively. Figure 4
shows the molecular structure of 5 with the atom num-
bering scheme, and Table 2 gives selected bond distances
and angles. The asymmetric unit of 7 consists of two
unique molecules; Figure 5 shows views of the two crys-
tallographically independent molecules (7.1 and 7.2)
identified in their numbering schemes by leading 1 and
2 characters, respectively. Both complexes are dinuclear,
and structural parameters for two equivalent halves of
the molecules, that lie either side of the NH unit of the
diphenylamine ring system, are reasonably similar. Very
weak intramolecular N3‐H3 … N2 and N3‐H3 … N2ˈ
hydrogen bonds are found in 5 and enclose S(6) rings,
whereas for 7 the intramolecular contacts are N13‐H13
… O12 and N13‐H13 … N14 for 7.1 but N23‐H23 … N22
and N23‐H23 … N24 for 7.2. The phenyl rings of the cen-
tral diphenylamine units are inclined to one another by
46.23(18)° in 5, and by 51.96(17)° and 50.94(16)° in the
two molecules of 7 with no suggestion of mirror symme-
try in these complexes. In each complex, the bis‐
hydrazone acts as a tetra‐anionic ligand and provides
two contiguous ONO tridentate domains that coordinate
the two R2Sn moieties. Coordination is clearly preceded
by keto‐amine to enol‐imine tautomerization of H4L,
accompanied by complete deprotonation of the resulting
four hydroxo substituents. The coordination spheres of
the two tin (IV) in each complex comprise the two C
atoms of methyl (5) or butyl (7) groups from the halo‐
FIGURE 4 The molecular structure of 5 showing the atom
numbering and with ellipsoids drawn at the 50% probability level.
Intramolecular hydrogen bonds are shown as dotted lines
alkyl starting materials, augmented by imine nitrogen,
phenolate and enolate oxygen atoms from the
deprotonated tridentate H4L ligand. With a coordination
number of five, the distinction between ideal square pyra-
midal or trigonal bipyramidal coordination geometries is
best made by calculating the trigonality index,[53]
τ5 = (α − β)/60, for each Sn (IV) center. In this equation,
α and β are the two largest bond angles around each tin
atom. The τ5 values for ideal trigonal bipyramidal and
square pyramidal geometry are zero and unity, respec-
tively. For complex 5, the τ5 values are 0.47 and 0.41 for
Sn1 and Sn2, respectively, suggesting a highly distorted
square pyramidal geometry with the N1 and N1′ imine
nitrogen atoms at the apices of the pyramid. The rigidity
of the chelate rings causes the structural distortion, and
the large covalent radius of the tin (IV) nuclei further
facilitate this deviation.[54,55] The corresponding indices
for the tin (IV) centers in the two unique molecules of 7
are 0.38 and 0.47 for Sn11 and Sn12 in 7.1, and 0.44 and
0.53 for Sn21 and Sn22 in 7.2. Thus, the coordination
environments of the Sn centers in 7.1 are distorted square
pyramidal similar to that found for complex 5. The atoms
at the apices are imine nitrogen (N11) for Sn11 and meth-
ylene carbon atom of one of the butyl groups (C137) for
Sn12. In contrast, the two Sn atoms in 7.2 adopt differing
coordination geometries, albeit only marginally so. Sn21
is in a distorted square pyramidal with the imine nitrogen
(N21) in the axial position comparable to the situation for
5, whereas Sn22 defies the trend and adopts highly
FIGURE 3 Crystal packing of 4 viewed along the b axis direction
with hydrogen bonds drawn as dashed lines

YOUSEFI ET AL.
9 of 15
FIGURE 5 The two unique molecules in the asymmetric unit of 7 showing the atom numbering with ellipsoids drawn at the 50%
probability level. Intramolecular hydrogen bonds are drawn as dotted lines
distorted trigonal bipyramidal geometry with the O23 and
O24 atoms in axial positions.
These sheets are stacked along the b axis direction by
Cg5 … Cg7x and Cg 6 … Cg8x offset π … π stacking inter-
actions with centroid to centroid distances both of
3.829(4) Å (Cg5, Cg6, Cg7 and Cg8 are the centroids of
the C1‐C6, C9‐C14, C1ˈ‐C6ˈ, C9ˈ‐C14ˈ rings, respectively,
with x = 2 − x, −y, 1 −z; Figure 6a). For 7, C‐H … Br
and C‐H … O hydrogen bonds combine with an extensive
series of C‐H … π (ring) contacts (Table S1) to stack the
complexes in sheets along the b axis direction (Figure 6
b). The structure is further stabilized by Cg7 … Cg16y
and Cg8 … Cg15y offset π … π stacking interactions with
centroid to centroid distances of 3.969(4) Å and 3.718(4)
Å (Cg7, Cg8, Cg15 and Cg16 are the centroids of the
C215‐C220, C223‐C228, C123‐C128 and C115‐C1120
rings, respectively, with y = 1 + x, 1 + y, z).
The Sn − N and Sn − O bond lengths in all complexes
(Table 2) are very close to the sum of the covalent radii of
the elements (Sn − N, 2.15 Ǻ and Sn − O, 2.10 Ǻ), and are
considerably shorter than the sum of van der Waals radii
of the corresponding atoms, 3.85 and 3.80 Ǻ, respec-
tively.[56,57] This suggests that the coordinate bonds are
strong in both complexes.
The methyl and n‐butyl substituents on the Sn atoms
of 5 and 7 have a significant influence on the structures
of these molecules, as might be expected given the signif-
icant differences in the steric requirements of these sub-
stituents. The most obvious result of varying the alkyl
substituents on the Sn atoms lies in the coordination
geometries that they adopt. Both Sn atoms of 5 adopt
heavily distorted square pyramidal with quite similar τ5
indices. However, only one of the two discrete molecules
of 7 also has both Sn atoms in a distorted square pyrami-
dal configuration, although the degree of distortion is
considerably less for Sn11. Furthermore, in the second
molecule Sn22 is trigonal bipyramidal while Sn21
remains square pyramidal. Clearly the necessity to
accommodate the much larger n‐butyl substituent is the
key determinant of these differences. Comparison of the
bond lengths and angles associated with the alkyl ligands
(Table S1) also points to a significant increase in steric
strain associated with the n‐butyl substituents. The Sn‐C
bond distances in both molecules of 7 are generally lon-
ger than those to the methyl groups, and the C‐Sn‐C
angles are either much greater or significantly less that
the corresponding parameters for 5. Finally, the extreme
variations in the Sn‐C‐C and C‐C‐C angles in the all of
the Sn‐CH2‐CH2‐CH2‐CH3 chains further attest to steric
strain in these systems.
3.2 | Spectroscopic studies
In the FT‐IR spectrum of the complexes, the band attrib-
uted to the C=O stretching vibration in the free ligand is
absent, confirming that the ligand coordinates in the
enolate form. The vibrational frequency of the C=N bond
is shifted very slightly to lower wavenumbers in the spec-
tra of the complexes, indicating that the azomethine
nitrogen atom is involved in coordination to the tin.
Appearance of the new bands in the ~400–600 cm−1
range attributed to ν (Sn‐N) and ν (Sn‐O) further confirms
coordination of nitrogen and oxygen to the metal.
1
Details of the H‐NMR spectral data of H4L and its
complexes in DMSO are presented in the Experimental
section. It is noteworthy that H4L is a symmetrically
substituted diphenylamine, and so the two chemically
equivalent sets of protons in the molecule are also mag-
netically equivalence in solution. Therefore one signal is
observed for each of the Ar‐OH, –NH‐N = and CH=N
protons. The ratio of the integrals of the alkyl/aryl reso-
nances of the R2Sn moieties to those of the ligand
In the crystal of 5, C‐H … O hydrogen bonds (Table S2)
link complexes into rows along a. C‐H … Br hydrogen
bonds link adjacent rows into sheets in the ac plane.

10 of 15
YOUSEFI ET AL.
FIGURE 6 Crystal packing of (a) 5 and (b) 7, viewed along the b axis. Hydrogen bonds are drawn as dashed lines and representative π … π
contacts are shown as dotted green lines. Ring centroids are shown as colored spheres
confirms the 2:1 coordination ratio. Signals due to the
phenol and the amide functions of H4L are absent in
the H‐NMR spectra of complexes, as expected with the
centers in complexes. The 119Sn chemical shift is highly
dependent on the coordination number, and an increase
in coordination number leads to a large upfield shift.
On the basis of the range reported empirically in the liter-
ature for organotin (IV) complexes, the complexes con-
tain 6‐coordinated tin atoms.[59–63] Because DMSO is a
coordinating solvent, the signal shifts to the region corre-
sponding to hexacoordinated species.[61] Unfortunately,
the solubility of all complexes was very low in CDCl3 or
other non‐coordinating solvents. For the diphenyltin
complex, the chemical shift appears at a higher field than
those with aliphatic substituents. This difference is unre-
markable that appears also in the starting materials,
Me2SnCl2 (+137 ppm), BuSn2Cl2(+122 ppm) and
Ph2SnCl2 (−27 ppm).[64] It has been suggested that this
trend is due to anisotropic shielding effects and possible
π‐interactions in the phenyltin compounds.[60]
1
complete deprotonation of the enol form followed by
coordination to tin. A singlet resonance attributable to
the two imine protons is accompanied by 119Sn satellites
in the spectra of all complexes. This is an evidence that
the two imine nitrogen atoms are chemically equivalent
in solution and both are coordinated to tin (IV) centers.
1
The H‐NMR spectrum of dimethyltin complex shows a
singlet at 0.69 ppm for SnMe2 protons flanked by satel-
2
lites due to J(119Sn‐1H) (Figure S1). The appearance of
only one signal for all 12 methyl protons is an evidence
that the two SnMe2 moieties have an equivalent chemical
2
environment in solution. Substitution of the J(119Sn‐1H)
values into the Lockhart–Manders equation[58] gives a
value of 145.1° for the CH3‐Sn‐CH3 angle in DMSO solu-
tion. The corresponding values from the X‐ray data were
126.6° and 129.7° for the two discrete tin centers, a dis-
crepancy probably due to variations between the solid
state and solution structures. Greater flexibility is likely
in solution with DMSO as a coordinating solvent that
increases the coordination number of tin to six.
3.3 | DNA‐binding
In order to explore the interaction between ligand and
complexes with CT‐DNA, the absorption spectral titration
with CT‐DNA under physiological conditions was used.
The results for the diphenyl complex have been given in
Figure 7, and for others are shown in Figures S2–S4. A
substrate such as a metal complex can interact with
DNA via a covalent and/or non‐covalent manner on the
basis of their geometry, size, hydrophobicity and
hydrogen‐bonding ability. In the covalent interaction, a
nitrogen donor base in DNA strands can be coordinated
to the metal. The non‐covalent DNA interactions include
three modes of binding: (a) intercalation; (b) groove (sur-
face); and (c) electrostatic. Important features of the sub-
strate, such as geometry, size, hydrophobicity and
1
The H‐NMR spectrum of 7 shows high‐field multi-
2
plets for the SnBu2 protons, but the J(119Sn‐1H) value
for Sn‐CH2 cannot be extracted from this spectrum
because of the complexity of the methylene multiplets.
All of the complexes show multiplets at 8.2–6.8 ppm for
aromatic protons of ligand. The phenyl protons of the
Ph2Sn groups in 6 also appear as multiplets in the range
7.63–7.60 ppm for the ortho protons with 3J(119Sn‐1H) sat-
ellites, and 7.25–7.28 ppm for meta and para protons.
The 119Sn‐NMR spectra of three complexes in
DMSO‐d6 show only single sharp up‐field signals showing
equivalent chemical environments for the pairs of tin

YOUSEFI ET AL.
11 of 15
FIGURE 7 Electronic absorption spectra for titration of 6 with increasing amounts of calf thymus (CT)‐DNA, 0, 0.5, 2, 5, 6 and 8 μg.
Arrows show the spectra changing upon the increase of CT‐DNA concentration. Inset: plots of –[DNA]/(εa − εf) vs. [DNA]
hydrogen‐bonding ability, can all contribute to the DNA‐
binding affinity. In this way, the presence of suitable
groups for the formation of hydrogen bonding, or the
presence of aromatic groups for hydrophobic interactions
with DNA and the existence or creation of the free coor-
dination sites in metal complexes can facilitate the inter-
ference with DNA double‐helix and lead to a potential for
interaction with DNA. Interaction mode leads to consid-
erable changes in the photophysical properties of com-
(bathochromism). These observations confirm the inter-
action of compounds with CT‐DNA, and hypochromism
and bathochromism in UV–Vis spectra are related to
the binding of the compounds to DNA through intercala-
tion arising from the interactions between the aromatic
parts of compounds and the π orbitals of DNA base‐pairs.
A large degree of hypochromism and red‐shift indicates
strong intercalative interaction.
To quantify the extent of binding of compounds to CT‐
DNA, the intrinsic binding constant, Kb, was calculated
by monitoring the changes in the absorbance at the corre-
sponding λmax with increasing concentrations using the
Wolfe–Shimer equation:[70]
pounds, and can be analyzed by
a number of
techniques. One of the methods that has widely been per-
formed to check DNA‐binding is monitoring the absorp-
tion spectra of compounds by adding a constant amount
of compounds to an increasing amount of DNA. The
spectral characteristics provide an evidence for the bind-
ing of the compounds to DNA. Although the binding
mode cannot be determined with certainty, nevertheless
binding of a metal complex to DNA through intercalation
may result in hypochromism and bathochromism in UV–
Vis spectra. Depending on the extent of red‐shift and
hypochromism, the strength of the intercalative interac-
tion can be realized. However, hyperchromism is associ-
½DNAꢀ
½DNAꢀ
1
¼
þ
ðεa − εbÞ ðεb − εfÞ Kbðεb − εfÞ
where [DNA] is the concentration of CT‐DNA, and εa, εb
and εf are the extinction coefficients for the apparent,
bound, and free ligand or organotin (IV) complex, respec-
tively. The ratio of the slope to the y‐intercept in the plot
of [DNA]/(εa – εf) vs. [DNA] gives the values of Kb (inset
ated with
a
non‐intercalative or an electrostatic
in Figure 8). The binding constants were calculated to be
−1
interaction with DNA.[14,65–69]
1.25 ×105, 1.56 × 105, 5.61 ×105 and 1.47 ×105
M
for 4–
In this way, the π‐π* transition of the ligand at about
300 nm and the related intra‐ligand transition in the com-
plexes were considered for the absorption spectral titra-
tion with CT‐DNA. All compounds exhibit similar
behavior with increasing amounts of DNA, which
includes the reducing of adsorption intensity at λmax
7, respectively. Comparing these values with the litera-
ture suggests that the interactions of compounds with
CT‐DNA are significant.[14,65–68,71–73] The binding con-
stants indicate that 6 has significantly higher affinity to
DNA. Possibly, the presence of more aromatic groups in
diphenyltin (IV) complex facilitates interacting with
DNA strands. Because the aromatic phenyl groups have
(hypochromism)
accompanied
with
red‐shift

12 of 15
YOUSEFI ET AL.
FIGURE 8 Agarose gel electrophoresis patterns for cleavage of plasmid DNA by ligand and complexes
hydrophobic interaction with the bases that make up the
DNA structure, therefore they have higher intercalation
between DNA base‐pairs and show a stronger DNA‐
binding effect.
with DNA amplification. In this assay, increasing
amounts of ligand and its organotin complexes with
genomic DNA for 1 hr at room temperature were treated.
For amplification of DNA, the PCR reaction mixture was
added on a thermal cycler. After 35 cycles, the PCR prod-
uct was electrophoresed on agarose gel followed by
ethidium bromide staining and UV visualization.
3.4 | Plasmid DNA cleavage
The results illustrate that all the compounds affected
on amplification of DNA and organotin complexes are
more effective than ligand, especially in 25 and 100 μM
concentration of compounds (Figure 9).
The DNA‐cleavage ability of ligand and its complexes was
studied by the agarose gel electrophoresis technique
using the DNA plasmid from the E. coli bacteria. The
electrophotogram for the cleavage activity of 4–7 is given
in Figure 8. No cleavage of DNA was observed with
ligand and complexes, but a slow migration in the band
of the plasmid could be due to change of the superhelix
form of the plasmid to relax form or linear form due to
intercalation of molecules into DNA strands.[67,68]
3.6 | Cytotoxicity
The in vitro cytotoxicity of the synthesized compounds
against MCF7 the human breast cancer line was deter-
mined using the MTT assay, in which mitochondrial
dehydrogenase activity was measured as an indication
of cell viability. The cytotoxicity of the compounds was
evaluated by curves obtained for each compound
(Figure 10). Based on the inhibition curve, the activity
of dibutyl complex is higher than the other compounds,
3.5 | Polymerase chain reaction‐based
assay
Polymerase chain reaction assays were performed to
investigate whether synthesized compounds interfere
FIGURE 9 Polymerase chain reaction
(PCR) inhibition assay with compounds

YOUSEFI ET AL.
13 of 15
ligand was coordinated to two diorganotin (IV) moieties
through two ONO tridentate domains: the enolic oxygen,
imine nitrogen and phenolic oxygen atoms. Both tin
atoms in all complexes have a similar environment with
a coordination number of five. The synthesized com-
pounds significantly interact with CT‐DNA mainly
through intercalation, which is affected on amplification
of genomic DNA and show cytotoxicity against the
human breast cancer line (MCF7).
FIGURE 10 The inhibition effects of compounds 4–7 on the
breast cancer cell line (MCF7) at different concentrations
ACKNOWLEDGEMENT
Support and funding of this work by Shahid Chamran
University of Ahvaz, Ahvaz, Iran (Grant no. 1396) is
gratefully acknowledged. The authors also thank the Uni-
versity of Otago for purchase of the diffractometer, and
the Chemistry Department, University of Otago for sup-
port of the work of JS.
so we are able to report IC50 values only for the butyl
derivative (IC50 = 37.5 μM). All of the cancer cells were
subjected to cell death after 72 hr of toxicity testing com-
pounds. The greater activity of di(n‐butyl)tin (IV) com-
plex compared with the dimethyl and diphenyl
complexes is in agreement with previous research results,
and can be described on the basis of the optimal balance
between the cytotoxicity, solubility and lypophilicity.[74–
SUPPLEMENTARY MATERIAL
78]
A survey of the anticancer activity of organotin (IV)
compounds during the last 10 years[79] shows that com-
plex 7, in comparison to cis‐platin and many similar
organotins, has good activity against the MCF‐7 cell line.
It is noteworthy that while the phenyl derivative has a
strong DNA‐binding effect, it shows a lower cytotoxicity
effect than butyl derivative and free ligand. In fact, not
all of the complexes with high cytotoxicity have better
binding activity with DNA.[80] Each substance can have
a toxic effect on the cell by various mechanisms, such as
destruction of cell membranes, prevention of protein syn-
thesis, DNA interaction, effect on certain genes or path-
ways involving cell survival. Therefore, a substance may
have a strong DNA‐binding effect, but it does not neces-
sarily have a strong pernicious effect on the cell, and
the other substance can have a cell death effect by bind-
ing to certain proteins without having a strong binding
with the DNA. On the other hand, the ability to penetrate
the cell membrane is an important factor in affecting the
intracellular components, and less inhibition of a com-
pound may be attributed to the low permeability across
the cell wall. The activity of ligand that is more than
dimethyl and diphenyl complexes may be attributed to
OH, NH and the imine groups, which interact with active
biological centers.[18,81,82]
CCDC 1852663, 1852664 and 1852665 contain the supple-
mentary crystallographic data for H4L·H2O, [Me4Sn2L]
and [Bu4Sn2L], respectively. These data can be obtained
free of charge via http://www.ccdc.cam.ac.uk/conts/
retrieving.html, or from the Cambridge Crystallographic
Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK;
fax: +44 1223 336 033; or e‐mail: deposit@ccdc.cam.ac.uk.
Some bond distances and angles for 5 and 7 (Table S1),
1
hydrogen bonding parameters (Table S1), H‐NMR spec-
trum of 5 (Figure S1), and electronic absorption spectra
for titration of 4, 5 and 7 with CT‐DNA (Figures S2–S4)
have been given.
ORCID
Tahereh Sedaghat https://orcid.org/0000-0002-3352-1932
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Three new dinuclear organotin (IV) complexes with
diphenylamine as linker have been synthesized and fully
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Simpson J, Shafiei M. Bis‐aroylhydrazone based on
2,2′‐bis substituted diphenylamine for synthesis of
new binuclear organotin (IV) complexes:
Spectroscopic characterization, crystal structures,
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