Synthesis, characterization, biological screenings and molecular docking study of Organotin(IV) derivatives of 2,4-dichlorophenoxyacetic acid

Article abstract of DOI:10.1016/j.molstruc.2018.11.011

New tri- and diorganotin (IV) derivatives of 2,4-dichlorophenoxyacetic acid with general formula: R₃SnL and R₂SnL₂: {Me₃SnL (1), Bu₃SnL (2), Me₂SnL₂ (3), Bu₂SnL₂ (4) and Oct₂SnL₂ (5), L = 2,4-dichlorophenoxyacetate} have been synthesized and characterized in solid state by elemental and FT-IR analysis, whereas in solution state by ¹H and ¹³C NMR spectroscopy. Compound 1 was also characterized by single crystal X-ray crystallography. The FT-IR data of compounds 1–5 confirm the bidentate binding mode of ligand with penta and hexa-coordinated arrangements around the Sn(IV) centre in solid state. The value of C–Sn–C angle for complexes 1 and 3 calculated from NMR (¹H and ¹³C) data using Lockart's equation were 114.7° and 114.9°, respectively which falls in the range of 5-coordinated geometry. The DNA binding of synthesized compounds were studied via UV–Vis spectroscopy and viscometry resulting in an intercalative mode of interaction. Molecular docking analysis of the studied compounds also supports the results of the UV–vis and viscometry. Moreover, interaction of the synthesized compounds with a cationic surfactant i.e., cetyltrimethyl ammonium bromide (CTAB) has been studied by conductometric method. Enzyme inhibition activity against α-amylase and α-glucosidase was carried out and compound 3 was found to possess maximum inhibition (88.1% and 91.3%, respectively). The theoretical study also enforce the experimental data for enzyme inhibition of the compound 3 (docking score = ?12.4096) by forming seven hydrogen bonds and two pi-H linkages with the Glu 276, Ala 278, Phe 300, Arg 312, Tyr 313, Asp 349, Asn 412, Phe 430 and Arg 439 residues of the binding pocket of the α-glucosidase. The potency of the compound 3 might be due to the presence of the strong electron withdrawing chloro group. IC₅₀ value of the brine shrimp activity revealed that triorganotin (IV) derivatives (1 and 2) were more toxic than their diorganotin (IV) analogues. Moreover, compound 2 has the MIC values of 12.5 μg/mL and 6.25 μg/mL against S. Aureus and M. Leuteus bacterial strains, respectively.

Full text of DOI:10.1016/j.molstruc.2018.11.011

Accepted Manuscript
Synthesis, Characterization, Biological Screenings and Molecular Docking Study
of Organotin(IV) Derivatives of 2,4-Dichlorophenoxyacetic Acid
Nida Naz, Muhammad Sirajuddin, Ali Haider, Syed Mustansar Abbas, Saqib Ali,
Abdul Wadood, Mehreen Ghufran, Gauhar Rehman, Bushra Mirza
PII:
S0022-2860(18)31315-2
10.1016/j.molstruc.2018.11.011
MOLSTR 25840
DOI:
Reference:
To appear in:
Journal of Molecular Structure
Received Date:
Accepted Date:
16 October 2018
05 November 2018
Please cite this article as: Nida Naz, Muhammad Sirajuddin, Ali Haider, Syed Mustansar Abbas,
Saqib Ali, Abdul Wadood, Mehreen Ghufran, Gauhar Rehman, Bushra Mirza, Synthesis,
Characterization, Biological Screenings and Molecular Docking Study of Organotin(IV) Derivatives
of 2,4-Dichlorophenoxyacetic Acid, Journal of Molecular Structure (2018), doi: 10.1016/j.molstruc.
2018.11.011
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Graphical Abstract: Pictogram
Synopsis for Graphical Abstract:
The novel organotin(IV) Carboxylates complexes interact with DNA via intercalative mode. Also they have strong tendency to
interact with CTAB (surfactant) as indicated by higher CMC values and the resulting in micelle formation plays an important role in
drug devilry. They show strong inhibitory potential against α-amylase and α-glucosidase. The DNA interaction and enzyme inhibition
by Molecular Docking strongly support the experimental data.

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Synthesis, Characterization, Biological Screenings and Molecular Docking Study of
Organotin(IV) Derivatives of 2,4-Dichlorophenoxyacetic Acid
a§
b*§
a*
c,d
a
Nida Naz , Muhammad Sirajuddin , Ali Haider , Syed Mustansar Abbas , Saqib Ali ,
e
e
f
f
Abdul Wadood , Mehreen Ghufran , Gauhar Rehman , Bushra Mirza
aDepartment of Chemistry, Quaid-i-Azam University Islamabad, 45320, Pakistan
bDepartment of Chemistry, University of Science and Technology Bannu, 28100, Pakistan
c
Nanoscience & Technology Department, National Centre for Physics, Islamabad, Pakistan
d
Department of Energy and Materials Engineering, Dongguk University, 30, Pildong-ro 1 gil,
Jung-gu, Seoul 100-715, Republic of Korea
eDepartment of Biochemistry, Computational Medicinal Chemistry Laboratory, UCSS, Abdul
Wali Khan University Mardan, Pakistan
fDepartment of Biochemistry, Quaid-i-Azam University Islamabad, 45320, Pakistan
§Authors Nida Naz and Muhmammad Sirajuddin Contributed equally
*Corresponding authors, e-mail: m.siraj09@yahoo.com (M. Sirajuddin); ahaider@qau.edu.pk
(A. Haider)
Abstract
New tri- and diorganotin(IV) derivatives of 2,4-dichlorophenoxyacetic acid with general
formula: R SnL and R SnL : {Me SnL (1), Bu SnL (2), Me SnL (3), Bu SnL (4) and Oct SnL
2
3
2
2
3
3
2
2
2
2
2
(5), L = 2,4-dichlorophenoxyacetate} have been synthesized and characterized in solid state by
1
13
elemental and FT-IR analysis, whereas in solution state by H- and C-NMR spectroscopy.
Compound 1 was also characterized by single crystal X-ray crystallography. The FT-IR data of
compounds 1-5 confirm the bidentate binding mode of ligand with penta and hexa-coordinated
arrangements around the Sn(IV) centre in solid state. The value of C-Sn-C angle for complexes 1
1
13
and 3 calculated from NMR ( H and C) data using Lockart’s equation were 114.7° and 114.9°,
respectively which falls in the range of 5-coordinated geometry. The DNA binding of
synthesized compounds were studied via UV-Vis spectroscopy and viscometry resulting in an
intercalative mode of interaction. Molecular docking analysis of the studied compounds also
supports the results of the UV-vis and viscometry. Moreover, interaction of the synthesized
compounds with a cationic surfactant i.e., cetyltrimethyl ammonium bromide (CTAB) has been
studied by conductometric method. Enzyme inhibition activity against α-amylase and α-

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glucosidase was carried out and compound 3 was found to possess maximum inhibition (88.1%
and 91.3%, respectively). The theoretical study also enforce the experimental data for enzyme
inhibition of the compound 3 (docking score = -12.4096) by forming seven hydrogen bonds and
two pi-H linkages with the Glu 276, Ala 278, Phe 300, Arg 312, Tyr 313, Asp 349, Asn 412, Phe
4
30 and Arg 439 residues of the binding pocket of the α-glucosidase. The potency of the
compound 3 might be due to the presence of the strong electron withdrawing chloro group. IC50
value of the brine shrimp activity revealed that triorganotin(IV) derivatives (1 and 2) were more
toxic than their diorganotin(IV) analogues. Moreover, compound 2 has the MIC values of 12.5
µg/mL and 6.25 µg/mL against S. Aureus and M. Leuteus bacterial strains, respectively.
Keywords: Organotin(IV) derivative; DNA interaction study; Cetyltrimethyl ammonium
bromide (CTAB); Enzyme inhibition activity; Molecular docking
1
.
Introduction
-1
Organotins are intensively used organometallic compounds worldwide (~ 50,000 t yr ) with
applications in the stabilization of plastics, precursors in glass coating and as antifungal agents in
textiles and other household’s items [1]. From the 1950s-2001, tributyltin (TBT), dibutyltin
(DBT), monobutyltin (MBT) and triphenyltin (TPhT) compounds are the most widely used
active ingredients in marine antifouling coating to resist the settlement of biofouling agents [2].
Despite the environmental and toxicological problems associated with the technological use of
organotin compounds [3], they are among the most widely employed organometallic species and
continue to surprise experts in the field of medicinal inorganic chemistry [4, 5].
Organotin compounds of the general formula R SnX display a wide range of biological effects
n
4-n
depending on the ligand X, its number n and on the type of the organic group R bound to the
metal ion [6, 7]. The organotin(IV) compounds have been widely investigated during the last few
decades because of their potential antibacterial and anticancer activities [8]. It is noticeable that
some organotin(IV) complexes showed higher in vitro cytotoxic activities than cis-platin [9].
Therefore, the synthesis of novel organotin complexes and the investigation of their biological
activities are of great importance [10].
Among the organotin compounds, organotin carboxylates are important from for biological point
of view as they possess broad applications in medicine [11]. Many of the diorganotin and
triorganotin compounds possess intriguing molecular structures such as ladder, O-capped cube,

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butterfly, drum, football cage, cyclic trimer and so on that resulted in promising applications.
The structure of organotin carboxylates can be tuned by the ligand type, reaction conditions and
the organotin groups [12]. In order to understand the regulation of synthesis and the structural-
biomedical application relation much attention has been paid on the preparation, characterization
and biological activities of organotin carboxylates against tumour cells, fungi, bacteria and other
microorganisms [13,14]. Nowadays, resistance of microorganisms and toxicity are some of the
major problems concerning the clinical use of drugs. Therefore, the search for new metal-
containing antimicrobial agents, more bio-specific and less toxic to the host and to the
environment, is particularly important. In the continuation of our work on organotin(IV)
carboxylates, herein, we report on the synthesis and spectroscopic studies of di- and
triorganotin(IV) derivatives of 2,4-dichlorophenoxyacetic acid. These complexes are investigated
for their DNA binding and screened for enzyme inhibition and microbial activity. Moreover, we
have studied their interactions with cationic surfactant, i.e., CTAB by conductometric method.
2
2
.
Experimental
.1.
Materials and methods
The organotin(IV) chlorides and oxide were purchased from Aldrich, USA and were used
without further purification. The solvents used were procured from E. Merck, Germany and dried
before use by the reported method [15]. The melting points were measured in a capillary tube by
using the Gallenkamp (UK) electro thermal melting point apparatus. The FT-IR spectra were
−
1
recorded on a Thermo Nicolet-6700 FT-IR Spectrophotometer in the range of 4000-400 cm .
1
13
The H- and C-NMR were recorded at room temperature in deuterated solvents on a Bruker
Advance Digital 300 MHz FT-NMR spectrometer. Elemental analyses were done using a CE-
440 Elemental Analyzer (Exeter Analytical, Inc). DNA binding studies were performed using
Ubbelohde viscometer and double beam spectrophotometer UV-1601 from Shimadzu, Japan.
The interaction of the synthesized compounds with CTAB was studied using ELICO (type CM
-1
8
2T) bridge equipped with platinized electrodes (cell constant = 1.02 cm ) conductometer.
.2. General procedure for the Synthesis of NaL and Organotin(IV) derivatives
2
The NaL was prepared by stirring a mixture of equimolar amount of sodium hydrogen carbonate
dissolved in distilled water) with 2,4-dichlorophenoxyacetic acid (suspended in distilled water).
(

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The mixture was stirred for 30 minutes to get the clear solution. The solvent was rotary
evaporated to get the desired product (Scheme 1) [16].
Triorganotin(IV) derivatives (1 and 2) were synthesized by refluxing the mixture of equimolar
NaL (5 mmol) and R SnL {R = CH (1), n-C H (2)} (5 mmol) in dried toluene for 6-7 h
3
3
4
9
(
scheme 1) [16]. It was then cooled to room temperature and filtered. The filtrate was rotary
evaporated to obtain the desired product which was recrystallized in chloroform and n-hexane
4:1 ratio).
(
Diorganotin(IV) derivatives (3 and 4) were synthesized by the same procedure [16] as used for
the triorganotin(IV) derivatives with the only difference of molar ratio: NaL (10 mmol) and
R SnCl (5 mmol) (scheme 1).
2
2
Complex 5 was synthesized by refluxing the mixture of 2,4-dichlorophenoxyacetic acid (10
mmol) and (C H ) SnO (5 mmol) in dried toluene using the Dean and Stark apparatus, for 6-7 h
8
17 2
and water formed during the reaction was removed (scheme 1).

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Scheme 1: Synthetic pathway for NaL and its Organotin(IV) derivatives

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2
2
.2.
DNA interaction study
.2.1. UV–visible spectroscopy
SS-DNA (20 mg) was dissolved in deionized water (pH = 7.0) by stirring for overnight and
stored at 4°C. 20 mM Phosphate buffer (NaH PO –Na HPO , pH = 7.2) was prepared in distilled
2
4
2
4
water. A solution of SS-DNA in 20 mM Phosphate buffer gave a ratio of UV absorbance at 260
and 280 nm (A /A ) of 1.8 which confirms that DNA is free from proteins [17-20]. The molar
2
60
280
-1
-1
absorption coefficient 6600 M cm (260 nm) was used to find concentration of SS-DNA which
-4
was 1.4 х 10 M. The 1 mM complex was dissolved in 60% DMSO. Keeping the complex
concentration constant and the SS-DNA concentration varying the UV absorption titrations were
made. The SS-DNA solution of equal amount was added to the reference and complex to get rid
of absorbance of DNA itself. Before taking measurements compound-DNA solutions were
incubate for about 10 min at 25 °C. Using 1 cm path length cuvettes absorption spectra were
recorded [17-20].
2
.2.2. Viscosity measurements
Ubbelodhe viscometer was used to determine the viscosity at a room temperature using a digital
stopwatch. The flow time of sample was measured in triplicate and the average was taken. The
viscosity data were plotted as (η/ηo)1/3 vs. binding ratio (r) of [Compound]/[DNA]. Where η is
the relative viscosity of DNA in the presence of compounds, η is the relative viscosity of DNA
o
only. Viscosity values were calculated from the observed flow time of DNA containing solutions
corrected for the flow time of 20 mM phosphate buffer solution (pH 7.2) alone. The viscosity for
DNA in the presence and absence of the compound was calculated from the following equations
[21]:
t ‒ to
ηo = t ‒ to and η =
to
2
.3.
CTAB Interaction study
The interaction of synthesized compounds with CTAB was investigated by conductometric
method. The measurements were performed on an ELICO (type CM 82T) bridge equipped with
-1
platinized electrodes (cell constant =1.02 cm ). The conductivity runs were carried out by
adding concentrated CTAB stock solution progressively into the thermostated solvent
-6
-1
(
dimineralized double-distilled water of specific conductivity (1-2 × 10 Scm ) at 30 °C. The

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critical micellar concentration of the pure CTAB was obtained from the plots of specific
conductivity as a function of the CTAB concentration. The CMC values were taken from the
intersection of the two straight lines drawn before and after the intersection point versus
surfactant concentration plots [22].
2
2
.4.
In vitro bioactivities
.4.1. Brine shrimp cytotoxicity assay
Brine shrimp cytotoxicity assay was used to determine the toxicity of the compounds [23, 24].
−
1
Artemia salina (brine shrimp) eggs (Ocean Star Inc., USA) were hatched in seawater (34 gL ).
After 24 h, ten shrimps were transferred to each vial using Pasteur pipette. The compounds with
−
1
final concentrations of 400, 200, 100, 50, 10, and 5 μgmL were added and the volume was
raised up to 5 mL adding artificial seawater. Vincristine sulphate was used as positive control
and DMSO was used as negative control. The experiments were performed in triplicate and vials
were incubated under illumination at 28 °C. After 24 h, survivors were counted and IC (lethal
5
0
concentration) values were calculated by using the Finney software.
.4.2. Antibacterial activity
2
The in vitro antibacterial evaluation of the synthesized compounds was performed using the disc
diffusion method [25]. In experiment two gram positive: Staphylococcus aureus (ATCC 6538)
and Micrococcus luteus (ATCC 10240) and two gram negative: Escherichia coli (ATCC 15224)
and Enterobacter aerogens (ATCC 13048) were cultured in nutrient broth for 24h at 37 °C.
These cultured strains were used as inoculums (1%) to run the assay. Each bacterial strain was
added to the nutrient agar medium at 45 °C, poured into sterile Petri plates and allowed to
solidify. 5 mL of the test compound with final concentration of 200 mg/mL was poured on sterile
filter paper discs (4 mm) and placed on nutrient ager plates, respectively. Kanamycin and DMSO
were used as positive and negative controls, respectively on each plate. The assay was performed
in triplicate and the plates were incubated at 37 °C for 24–48 h. The antibacterial activity of the
compounds was determined by measuring the diameter of zones showing complete inhibition
(mm) with the help of vernier caliper.
2
.4.3. Antifungal assay
The in vitro antifungal evaluation of the synthesized compounds was performed using the disc
diffusion method [25] against the following fungal strains: Mucor species (FCBP 0300),

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Aspergillus niger (FCBP 0198) and Aspergillus fumigates (FCBP-066). Sabouraud dextrose agar
(SDA) was used to culture the fungal strains at 28°C for 5 to 7 days. Actively growing fungal
spores of each strain were spread with the help of autoclaved cotton swaps on solidified SDA
Petri plates under sterile conditions. The compounds (5 mL of each final concentration of 200
mg/mL) were transferred on sterile filter paper discs (4 mm) and placed on SDA plates.
Terbinafine and DMSO served as positive and negative controls, respectively on each plate. All
plates were incubated at 28 °C for 5 to 7 days and with the help of vernier caliper the fungal
growth was measured (mm).
2
.4.4. α-Amylase inhibition assay
The compounds were tested for their enzyme inhibition activity against α-amylase by the
previously reported method [26]. For assay 5 μL of each test compound with the final
concentration of 400 μg/mL was mixed with 40 μL of starch (0.05 %) and 30 μL of potassium
phosphate buffer (pH 6.8) in 96-well micro titer plates followed by the addition of 10 μL of α-
amylase enzyme (0.2 U/well). Acarbose and DMSO were used as positive and negative control,
respectively. The plates were incubated for 30 min at 50 °C and 20 μL HCl (1M) as stopping
reagent was added. Then 100 μL of iodine reagent (5 mM KI and 5 mM I ) was added to check
2
the presence and absence of starch and absorbance was measured at 540 nm with microplate
reader (Bio Tek, Elx800, USA). The experiments were performed in triplicate and percent
inhibition was calculated.
2
.4.5. α-Glucosidase inhibition assay
p-Nitrophenyl-α-D-glucopyranoside, Acarbose and Baker’s Yeast alpha glucosidase were
purchased from Sigma (USA). The alpha glucosidase was dissolved in 100 mM phosphate buffer
(pH 6.8) and was used as the enzyme extract. p-Nitrophenyl-α-D-glucopyranoside was used as
the substrate. Organotin compounds were used in the concentration of 400 μg/mL. Compounds
were mixed with 320 μL of 100 mM phosphate buffer (pH 6.8) at 30 °C for 5 minutes. Sodium
hydroxide (3 mL of 50 mM) was added to the mixture and the absorbance was read at 410 nm.
The control samples were prepared without any compound. Acarbose was used as the reference
alpha glucosidase inhibitor [27-29]. All tests were performed in triplicate. The percentage was
determined by mean of inhibitory values.

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2
.4.6. Molecular docking
Docking is a significant tool to explore the interactions between an inhibitor and the target [30].
To find the binding interactions of these compounds in the active sites DNA, α-glucosidase and
α-amylase, the MOE-Dock program was used to perform molecular docking. The crystal
structure of α-glucosidase is not available yet, so, we used homology model as described by
Ming Liu et al., [31] while the 3D crystal structure of the α-amylase (1HNY) and DNA (1BNA)
were retrieved from the Protein Databank (PDB). The structures of the derivatives were built in
MOE (www.chemcomp.com) and energy minimized using the default parameters of the MOE.
The DNA, α-glucosidase and α-amylase were allowed to dock to the compounds using MOE by
the default parameters i.e., Placement: Triangle Matcher, Rescoring: London dG. For each ligand
ten conformations were generated. The top-ranked conformation of each compound was used for
further analysis.
3
. Results and discussion
All the compounds were synthesized in good yield and their melting point along with other
physical data are given in Table 1.
Table 1: Physical data of the synthesized organotin(IV) derivatives 1-5a
Comp. No % Yield
M.P
M. Formula
M. Wt
Elemental analyses data (%)
Calculated (Found)
C
H
1
2
3
4
5
73
76
63
69
82
89-90
C H Cl O Sn 383.84
34.42 (34.02)
47.09 (47.87)
33.90 (33.71)
42.83 (42.25)
48.95 (49.01)
3.68 (3.87)
6.32 (6.99)
3.20 (3.11)
4.19 (4.25)
5.65 (5.55)
1
1
14
2
3
158-160 C H Cl O Sn 510.08
2
0
32
2
3
143-145 C H Cl O Sn 566.84
1
6
18
4
6
177-179 C H Cl O Sn
673
2
4
28
4
6
163-165 C H Cl O Sn 785.21
3
2
44
4
6
a)
Numbering of compounds is according to Scheme 1.
3
.1.
FT-IR spectra
From FT-IR data in Table 2, the absence of broad peak for acidic -OH, shifting of two C=O
bands (asymmetric and symmetric) and appearance of two new peaks for Sn-C and Sn-O in the
-1
-1
range of 540-575 cm and 440-485 cm , respectively confirms the synthesis of complexes 1-5.

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According to Deacon and Phillips, Δν for monodentate bonding increases compared to the free
anion; for bidentate chelating Δν decreases; and, for bidentate bridging, Δν remains similar to the
free anion [32]. The values of Δν are calculated as: Δν = νCOOasym - νCOOsym and indicate the
-
bidendate binding mode of the COO moiety around the Sn atom in the present series of
compounds.
Table 2: FT-IR data ν (cm⁻¹) of HL, NaL and its organotin(IV) derivatives 1-5a
Comp. No ν(O-H)
νCOOasym
1667
νCOOsym ∆ν(νCOOasym- νCOO ) νSn-C νSn-O
sym
HL
3420
1421
1443
1347
1370
1374
1375
1379
246
195
161
153
135
131
134
-
-
NaL
-
-
-
-
-
-
1638
-
-
1
2
3
4
5
1508
546
542
563
566
535
439
487
437
479
453
1523
1509
1506
1513
a) Numbering of compounds is according to Scheme 1.
3
3
.2.
NMR spectroscopy
1
.2.1. H-NMR
Compounds 1 and 3 which contain the methyl protons bonded to tin metal exhibited a definite
2
119/117
1
singlet at 0.71 ppm and 0.89 ppm having J (
Sn- H) values of 65 and 98 Hz, respectively.
2
The J values fall in the range of the 5-and 6-coordinated trigonal bipyradial and octahedral
geometries around the tin atom for tri- and di-organotin(IV) derivatives, respectively [4]. In
compounds 2, 4 and 5 the terminal methyl protons of butyl and octyl groups bonded to tin atom
gave a triplet each whereas all the rest of protons show multiplets. All the proton signals for
ligand are observed at same positions after complexation except those which are close to the
COO group show slightly downfield shift because of ligand to metal charge transfer which
1
further confirmed the complex formation. The H NMR values of all the synthesized complexes
are given in the Table 3.

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1
a
Table 3: H NMR data (ppm) of organotin(IV) derivatives 1-5
Comp.
No
OH (s, H2 (s, 2H) H4 (d, 1H) H5 (d, 1H)
1H)
H7 (s,
1H)
Sn-H
HL
11.01
-
4.26
4.66
6.86-6.82
6.75-6.72
7.27-7.23
7.18-7.15
7.48
-
1
2
3
7.31
7.39
7.41
0.71 (s, 9H, Hα),
2
1
119
J[ H- Sn] = 65 Hz
C-Sn-C = 114.9°
3H (t)0.84-0.8,
-
-
4.66
4.77
6.75-6.72
6.81-6.78
7.16-7.12
7.21-7.18
2
H (t)1.01-1.46
H (m)1.23-1.30
0.89 (s, 9H, Hα),
4
2
1
119
J[ H- Sn] = 98 Hz,
C-Sn-C = 145°
Sn-(C H )
4
5
-
-
4.77
4.76
6.78-6.75
6.78-6.75
7.19-7.15
7.19-7.15
7.42
7.42
4
9 2
0
.02-1.76 (m)
Sn-C H
8
17
0
.02-1.84 (m)
a) Numbering of compounds is according to Scheme 1.
3
.2.2. 13C NMR
All complexes have clearly been characterized by 13C-NMR, where the appearance of
characteristic peaks of alkyl groups around the tin centre that were absent in the free ligand
spectrum and the downfield shift of the peak of carbonyl carbon confirms the assimilation of
1
3
organotin moiety (Table 4). In C-NMR of compounds 1 and 3, the coupling constants for
1
119/117
13
satellites due to J[
Sn- C] coupling, have been calculated to be 432 Hz and 776 Hz,
respectively that corresponds to the θ(C-Sn-C) value of 114.7° and 144.8°, respectively by
applying Lockhartꞌs equation. The θ values suggest a 5-coordination geometry for
triorganotin(IV) derivatives and a 6-coordination geometry for diorganotin(IV) derivatives,
respectively in solution [33].

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1
3
a
Table 4: C NMR data of HL and its organotin(IV) derivatives 1-5
Comp. No
C1
C2
C3
C4
C5
C6
C7
C8
Sn-C
-
HL
1
170.5 68.6 154.2 115.6 127.1 127.9 129.2 122.2
172.6 66.6 152.0 114.1 126.4 127.4 130.2 123.9
-4.6 (Cα),
1
13
119
J[ C- Sn] =
32 Hz, C-Sn-C
114.7°
172.4 65.4 152.7 114.3 126.3 127.6 130.4 123.7 17.6 (C ), 25.4
4
=
2
3
α
(
C ), 24.7 (C ),
β γ
1
3.2 (Cδ)
175.1 66.0 152.1 114.5 127.4 128.5 130.5 124.2
1.05 (Cα),
1
13
119
J[ C- Sn] =
7
76 Hz, C-Sn-C
144.8°
177.2 66.3 152.3 114.4 127.0 127.4 130.5 124.2 18.6 (C ), 26.4
=
4
5
α
(
C ), 25.9
β
(
C ),13.6 (C )
γ δ
177.2 66.3 152.3 114.3 127.6 127.4 130.5 124.2 14.1-33.3 (Cα-δꞌ)
a) Numbering of compounds is according to Scheme 1.
3
.3.
Crystal structure of compound 1
Crystal structure of compound 1 is shown in Figure 1 whereas crystal data and structural
refinement parameters are given in Table 5. The selected bond lengths and bond angles are
reported in Table 6. The data reveals that the compound 1 adopted monoclinic crystal system
with P2 /c space group. The carboxylate ligand coordinates with two Sn(IV) atoms in a bridging
1
mode. The geometry around the Sn atom is distorted trigonal bipyramidal, wherein three methyl
groups are bonded to the Sn atom at equatorial positions with almost identical bond distances
[mean Sn-C = 2.113 Å] and two O atoms of the carboxylate ligand are bonded asymmetrically
with significantly different Sn-O distances of 2.198 Å and 2.468 Å [34]. The C-Sn-C bond
o
o
o
angles (C9-Sn1-C10, C10-Sn1-C11, C9-Sn1-C11) are 114.92 , 125.06 and 119.19 , respectively
o
o
and their sum is 359.17 . The O1-Sn1-O2 bond angle is 171.58 . The slight distortion is due to

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the asymmetrically bonded carboxylate ligands. The τ value (τ = (β-α)/60, where β is the largest
basal angle around the tin atom while α is the next largest angle around the tin atom) is 0.94 that
falls in the range of 5-coornidated distorted trigonal bipyramidal geometry [16, 34]. Packing
diagram of complex is shown in Figure 2.
Figure 1: The asymmetric unit of complex 1 with displacement ellipsoids drawn at the 50%
probability level. Symmetry codes: (i) -x, 1/2+y, 1/2-z; (ii) -x, -1/2+y, 1/2-z.

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Figure 2: Crystal packing structure of complex 1 Packing showing the formation of polymeric
one-dimensional chains parallel to the b axis. C-H...π interactions linking chains into layers
parallel to the (-1 0 2) plane are shown as dashed lines.

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Table 5: Crystal data and structure refinement for complex 1
Empirical formula
Formula weight
Temperature/K
Crystal system
Space group
a, b, c /Å
C H Cl O Sn
11 14 2 3
383.81
294(2)
monoclinic
P2 /c
1
10.6125(6), 10.4869(6), 13.5397(8)
α, β, γ /°
90, 104.0598(8), 90
Volume/Å3
1461.72(15)
Z
4
3
ρ g/cm
calc
1.744
μ/mm-1
2.107
F(000)
752.0
Crystal size/mm3
Radiation
0.320 × 0.280 × 0.200
MoKα (λ = 0.71073)
3.956 to 50.994
2
θ range for data collection/°
Index ranges
-12 ≤ h ≤ 12, -12 ≤ k ≤ 12, -16 ≤ l ≤ 16
15751
Reflections collected
Independent reflections
Data/restraints/parameters
Goodness-of-fit on F2
Final R indexes [I>=2σ (I)]
Final R indexes [all data]
Largest diff. peak/hole / e Å-3
CCDC
2711 [R = 0.0285, R
= 0.0183]
sigma
int
2711/0/157
1.122
R = 0.0230, wR = 0.0573
1
2
R = 0.0238, wR = 0.0577
1
2
0.27/-0.73
1823040
Table 5: Selected bond angles (°) and bond lengths (Å) for complex 1

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Selected bond angles
Atom Atom
Atom
Bond Angle Atom
Atom
Atom
Bond Angle
O1
C9
Sn1
Sn1
Sn1
Sn1
Sn1
Sn1
O2
O1
O2
O1
O2
C9
171.58(6)
88.99(10)
83.34(10)
95.02(10)
91.38(11)
114.92(15)
C11
C11
C11
C11
C1
Sn1
Sn1
Sn1
Sn1
O1
O1
O2
94.71(10)
86.06(10)
119.19(15)
125.06(14)
123.35(15)
148.25(16)
C9
C9
C10
C10
C10
C10
Sn1
Sn1
C1
O2
Selected Bond lengths
Atom Atom
Bond lengths (Å)
Atom
Atom
Bond lengths (Å)
Sn1
Sn1
Sn1
Sn1
Sn1
Cl1
Cl2
O1
O2
2.198(17)
2.463(18)
2.122(3)
2.110(3)
2.108(3)
1.740(3)
1.727(3)
O2
O2
O3
C1
C3
C3
O1
Sn1
C1
C3
C2
C4
C8
C1
2.463(18)
1.221(3)
1.376(3)
1.508(4)
1.381(4)
1.382(4)
1.277(3)
C9
C10
C11
C6
C8
3
3
.4.
DNA interaction studies
.4.1. UV-Visible spectroscopy
The absorption spectrum was initially used to study the interaction between the SS-DNA and the
synthesized complexes. Figures 3 and 4 represent the representative complexes interaction with
different concentrations of DNA. It was observed that the synthesized compounds show one
strong absorption peak in the range of 267-286 nm which is due to the π-π* transitions of
aromatic ring [35]. With the increasing concentrations of DNA, hypochromic along with minor
bathochromic shift was observed in the spectrum of the free compound peak which is the sign of
intercalation. In the intercalation mode of interaction, the drug insertion occurs between the two
strands of DNA which is generally adopted by the polycyclic or aromatic ring having planar
structures due to resemblance with DNA bases [35]. After the intercalation, the π* orbital of

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intercalated molecule interacts with the π orbital of the base pairs thus reduction of π-π*
transition occur resulting in bathochromic effect, which support the intercalative mode of
interaction. At λmax reduction in the absorbance appears as a result of π-π* transition which was
-1
used to measure DNA binding constant K (M ) from the intercept of graph plotted between
3
4
A /(A-A ) vs 1/[DNA]. High drug-DNA interaction in the range of 10 to 10 was found for all
o
o
the synthesized complexes. For di- and triorganotin(IV) complexes UV-Vis absorption studies
were carried out and the results of binding constant values (Table 6) confirm that
triorganotin(IV) derivatives show higher values of binding constants than their diorganotin(IV)
because of more lipophillic character of triorganotin(IV) derivatives. Based on the variation in
absorbance, the binding constant of this complex with DNA was determined according to
Benesi-Hildebrand equation [36].
A0
G
1
G



A A0 HG G
K

DNA

HG G
Where K is the binding constant, A and A are the absorbances of the drug and its complex with
o
DNA, respectively, and
and
are the absorption coefficients of the drug and the drug-
DNA complex, respectively. The association constant was obtained from the intercept-to-slope
ratios of A /(A-A ) vs. 1/[DNA] plot.
o
o
G
G
6
K = c/m х 10 (c = intercept =
and m = slope =
)
(for binding constant)
HG G
HG G
ΔG = -RT lnK (R = 8.314 J/mol, T = 298K)
(for Gibbꞌs Free energy, ΔG)

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Figure 3: Absorption spectrum of 2 mM of complex 1 in the absence and presence of 5 to 35 µM
of DNA. The arrow direction indicates increasing concentrations of DNA. The inset graph
-1
represents the plot of A /A-A vs. 1/[DNA] (µM) for the calculation of binding constant (K)
o
o
and Gibb’s free energy (∆G).

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Figure 4: Absorption spectrum of 2 mM of complex 5 in the absence and presence of 5 to 35 µM
of DNA. The arrow direction indicates increasing concentrations of DNA. The inset graph
-1
represents the plot of A /A-A vs. 1/[DNA] (µM) for the calculation of binding constant (K)
o
o
and Gibb’s free energy (∆G).
Table 6: Binding constant (K) and Gibbꞌs free energy (∆G) values calculated from UV-Vis.
for organotin(IV) derivatives 1-5a
-
1
∆G (kJ.Mol-1)
-24.13
Comp. No
K (M )
1.7 × 104
2.03 × 104
3.21 × 103
6.41 × 103
2.9 × 103
1
2
3
4
5
-24.57
-20.00
-21.72
-19.75
Numbering of compounds is according to Scheme 1.

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3
.4.2. Viscometry
Viscometric technique was used to further conform the binding mode of the synthesized
compounds with SS-DNA. The concentration of the compounds was kept constant while that of
1
/3
the DNA was varied. The relative viscosity (ɳ/ɳ₀) was plotted vs. concentration ratio of drug-
DNA adduct as shown in Figure 5 for the representative compounds. The intercalative mode was
favored when increase in viscosity was observed because when the drug interacts with DNA
bases the attractive forces between the DNA bases turn out to be weak and conformational
changes take place with increase in the torsional angle and finally the length of DNA increases
by uncoiling the two strand of DNA [37]. In terms of time flow the relative viscosity also
increases due to the increase in DNA length.
Figure 5: Effects of increasing amount of representative complexes 1 and 5 on relative viscosity
of DNA at 25±1 °C. [DNA] = 2.37×10⁻⁵ M.
3
.5.
Interaction with CTAB
The specific conductance of synthesized organotin (IV) complexes (1mM) with CTAB (0-0.007
M) were recorded to study the complex–CTAB interactions. The data of conductance were
plotted against the CTAB concentration used as shown in Figure 6 which shows that by
increasing the CTAB concentration the conductance increases and at 0.0039 M CTAB
concentration, an inflection point was observed which is the point at which CTAB monomers
combine to shape a structure called micelle. That inflection point is known as critical micelle
+
-
concentration (CMC) of CTAB. By adding the CTAB, the CTAB and Br ions were formed
which reduced the surface tension of aqueous media so increases the conductance [38]. The
conductance was observed to be sharply increase in the pre-micellar concentration region while

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the free monomers were used to form micelle in post-micellar concentration thus lowered the
speed of increase in conductance [39]. When the complex was present then the CTAB CMC
value was observed at high concentration as compared to its aqueous solution because the
+
complex molecules strongly bind to the CTAB monomers. On interaction of complex
+
molecules, the repulsion between CTAB monomers decreases by reducing its charge density.
ΔG is the free energy for the micellisation process which can be calculated by the equation
m
ΔG = 2.303(1+β) RTlogX
m
cmc
where β = the degree of the counter ion binding and
β = 1-α
α is the extent of dissociation or ionization which is calculated before and after CMC from the
ratio of plots.
α = S /S
2
1
cmc is taken in mole fractions called as Xcmc which in aqueous solution can be calculated as
Xcmc = cmc/55.55
ΔG and CMC for the complexes are given in Table 7, the variation of G values at highest
m
concentration and in the absence of surfactant is the ΔG and its negative value indicate
m
instinctive solubilization of compound in micelle because of attractive forces between the
compound and positively charged micelle results in stable CTAB-complex system. Specific
conductance data for the complexes in presence of CTAB (0–0.007M) were recorded to measure
complex–CTAB interaction parameters.
Table 7: CMC and Gibbꞌs free energy (∆G) values of selected organotin(IV) derivatives 1-
4a
-1
Comp. No
CMC value
∆G value (kJ.mol )
1
2
3
4
0.0039
0.0028
0.0015
0.0013
-38.5
-34.2
-39.4
-32.5
Numbering of compounds is according to Scheme 1.

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Figure 6: Specific conductivity curves of CTAB in the presence of the complex 1 in water
3
3
.7. In vitro Biological evaluation
.7.1. Antibacterial activity
All the synthesized compounds were screened for in vitro antibacterial activity against four
bacterial strains including two Gram positive (Staphylococcus aureus and Micrococcus luteus)
and two Gram negative strains (Enterobacter aerogenes and Escherichia coli) using agar disc
diffusion method. The synthesized compounds were dissolved in DMSO at concentration of 400
µg/mL. The results showed that most of the compounds were active against all the strains.
Compounds 2, 3 and 4 were most potent showing activity against most of the tested strains.
Further compound Bu SnL has shown outstanding activity against M. leuteus showing maximum
3
zone of inhibition 53.0 mm. Bu SnL compound was found active against all the tested bacterial
2
2
strains. The results of all the compounds are summarized in the Tables 8 and 9.

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Table 8: Antibacterial activity data of HL and its organotin(IV) derivatives 1-5a
Comp. No.
Zone of inhibition in mm (Conc. 400 µg/mL)
E. coli
E. aerogenes
S. aureus
M. leuteus
HL
-
12.4
6.3
7
-
-
-
-
1
-
-
2
-
-
7.4
-
53.0
11.0
11.4
-
3
4
9.1
-
12.5
-
9.4
-
5
Kanamycineb
21.2
22.4
23.1
24.6
a) Numbering of compounds is according to Scheme 1. b) Standard drug
Table 9: Minimum inhibitory concentration (MIC) of HL and its organotin(IV) derivatives
-5a
1
Comp. No.
MIC (µg/mL)
S. aureus
E. coli
-
E. aerogenes
M. leuteus
HL
1
-
-
-
200
25
-
200
12.5
200
50
-
-
2
200
-
6.25
100
100
-
3
100
50
4
200
-
5
-
a) Numbering of compounds is according to Scheme 1.
3
.7.2. Antifungal activity
The synthesized compounds were subjected to in vitro antifungal activity against three fungal
strains (Mucor species, A. niger and Aspergillus fumigatus) using the disk dilution method [29].
Turbinafine was used as the standered drug. All the compounds, except HL, 3 and 5 have shown
antifungal activity against the various strains. Compound 2 shows higher activity than the
reference drug against M. mucor. Data of the antifungal activity is shown in Table 10.

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Table 10: Zone of inhibition (mm) of antifungal activity of HL and its organotin(IV)
derivatives 1-5a
Comp. No
Zone of inhibition in mm (Conc. 400 µg/mL)
A. fumigates
A. niger
M. specie
HL
-
-
13.1
-
-
1
15.4
21.2
-
10.7
23.6
9.0
6.2
-
2
3
-
4
8.1
-
8.3
-
5
Terbinafineb
28.6
26.4
21.0
a) Numbering of compounds is according to Scheme 1. b) Standard drug
3
.7.3. Cytotoxicity
To screen out the biologically active compounds, brine shrimp cytotoxicity assay was performed
and results are summarized in Table 10. Brine shrimp cytotoxicity assay serves as predictive test
for the identification of bioactive compounds to evaluate anticancer potential of any compound.
−
1
All the tested compounds are highly cytotoxic with IC value as low as 0.69 휇gmL (Table 10).
5
0
It is important to note that these compounds are more potent than the vincristine sulphate
positive control) which means that these compounds have strong potential for the anticancer
(
screening. Compounds 1-4 had shown moderate level of lethality. Compound 5 was found totally
lethal against the shrimps.

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Table 10: Brine shrimp cytotoxicity data of HL and its organotin(IV) derivatives 1-5a
Comp.
No.
Percent mortality after 24 h at different concentrations (µg/mL)
IC50
400
70
200
55
100
50
50
40
10
35
60
50
40
45
25
5
HL
1
10
40
40
30
30
15
9.81
0.69
0.81
1.07
1.38
18.83
100
100
100
100
50
100
100
100
100
50
100
100
100
80
100
100
100
70
2
3
4
5
50
45
a) Numbering of compounds is according to Scheme 1.
3
.7.4. α-Amylase assay
The compounds were evaluated for their enzyme inhibition potential against α-amylase enzyme
and results in the form of percentage values as given in Table 12. Experiment was performed in
triplicates and acarbose was used as a positive control. The results showed that the only
compound 3 exhibited enzyme inhibition activity with percentage value of 88.1 and other
compounds were found inactive.
3
.7.5. α-Glucosidase inhibition activity
α-glucosidase inhibition assay was performed on all test compounds. Among the tested
compound all the compounds except compound 5 showed inhibition activity. Compounds 2 and
3
showed superb inhibition activity. Rest of the compounds showed moderate activity. Results of
alpha glucosidase inhibition assay are given in Table 12.
Table 12: % inhibition of alpha amylase and alpha glucosidase inhibition of HL and its
organotin(IV) derivatives 1-5a
Comp. No
α-amylase (% inhibition)
α-glucosidase (% inhibition)
HL
1
7.1
7.8
87.4
67.7
76.6
91.3
83.0
41.9
2
11.2
88.1
6.4
3
4
5
10.5

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a) Numbering of compounds is according to Scheme 1.
3
.8.
Docking analysis of DNA
To explore the binding mode of these derivatives, molecular docking was performed. The
docking results showed that all the metal-complex derivatives bind to DNA by intercalation,
which is the only interaction mode we obtained by docking. From the docking conformation of
the most active compound, compound 3 (docking score = -12.2325), it was observed that this
compound established two H-donor interactions with active site residues DC 9 and DG 10 of
DNA respectively as well as four pi-H linkages with the DT 7, DT 8, DC 9 and DG 10 residues
of the DNA (Figure 7). An excellent arrangement was obtained as the best docked pose showed
important binding features mostly based on interactions due to various interacting moieties of
derivatives, and phosphorus atom including its versatile structural features.
Figure 7: Docking orientation of 3 inside the active site of DNA.
3
.9.
Docking analysis of α-Glucosidase and α-Amylase
The docking results of the compounds with the α-amylase and α-glucosidase enzymes have
given good information about the nature of the binding mode that was excellent correlated with

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the experimental results. Compound 3 is the most active compound with highest docking score (-
1
0.8071) as well as inhibitory activity (88.1% inhibition). Compound 3 is bound to the α-
amylase in an adequate manner through one H-acceptor interaction and pi-pi interactions with
the active site residues Thr 163 and Trp 59 (Figure 8). The reason of the good activity of the
ligand 3 could be accredited to this strong binding with amino acid residues through polar
interaction as well as metal ligation. From the molecular docking studies, it was observed that
the top ranked conformation of all the compounds fit well inside the active site of the homology
model of α-glucosidase (Glu276, Asp 349 and Arg439) [40]. From the docking conformation of
the compounds, it was revealed that compound 3 (% inhibition 91.3, docking score = -12.4096)
formed seven hydrogen bonds and two pi-H linkages with the Glu 276, Ala 278, Phe 300, Arg
3
12, Tyr 313, Asp 349, Asn 412, Phe 430 and Arg 439 residues of the binding pocket of the α-
glucosidase as shown in Figure 9. The potency of the compound 3 might be due to the presence
of the strong electron withdrawing group (-Cl).
Figure 8: Docking conformation of compound 3 in the active site of α-amylase.

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Figure 9: Docking conformation of compound 3 in the active site of α-glucosidase.
Conclusions
New di- and triorganotin(IV) complexes of oxygen donor ligand have been synthesized in
quantitative yields. The appearance of new peaks for Sn-O and disappearance of O-H peaks of
acidic OH in the FT-IR spectra indicated the formation of organotin(IV) complexes.
1
13
Multinuclear NMR ( H and C) data further confirmed the formation of complexes. Compound
was also analyzed by X-ray single crystal analysis which revealed polymeric type bridged
1
trigonal bipyramidal structure with bidentate bridging mode of the carboxylate moiety. For SSD
DNA-Complex binding studies, the decrease in the absorbance (UV-Visible) and increase in the
relative viscosity supports the intercalative mode of interaction for the screened compounds.
Moreover, in case of complex-CTAB interactions, the higher value of CMC supports the idea of
high binding capability of organotin-withCTAB micelles due to electrostatic interations while
the negative value of energy changes suggests the spontaneous nature of transfer of organotin
molecules from bulk water to CTAB micelle. Biological studies of the compounds reveal that
triorganotin(IV) derivatives have higher activity than the corresponding diorganotin(IV), perhaps