Synthesis, characterization and DNA-intercalation studies of two ferrocene-based Fe-Sn heterobimetallic compounds, and crystal structure of trimethyltin (p-ferrocenyl)benzoate

Article abstract of DOI:10.1080/00958972.2017.1384547

In this article, we report the synthesis of two (A3 and A4) new ferrocene-based heterobimetallic compounds of Fe and Sn. Both compounds are structural isomers with formula (C₅H₅)Fe(C₅H₄C₆H₄

Full text of DOI:10.1080/00958972.2017.1384547

Journal of Coordination Chemistry
ISSN: 0095-8972 (Print) 1029-0389 (Online) Journal homepage: http://www.tandfonline.com/loi/gcoo20
Synthesis, characterization and DNA-
intercalation studies of two ferrocene-based Fe-Sn
heterobimetallic compounds, and crystal structure
of trimethyltin (p-ferrocenyl)benzoate
Ataf Ali Altaf, Nasir Khan, Bhajan Lal & Amin Badshah
To cite this article: Ataf Ali Altaf, Nasir Khan, Bhajan Lal & Amin Badshah (2017): Synthesis,
characterization and DNA-intercalation studies of two ferrocene-based Fe-Sn heterobimetallic
compounds, and crystal structure of trimethyltin (p-ferrocenyl)benzoate, Journal of Coordination
Chemistry, DOI: 10.1080/00958972.2017.1384547
To link to this article: http://dx.doi.org/10.1080/00958972.2017.1384547
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Date: 26 September 2017, At: 05:14

Publisher: Taylor & Francis
Journal: Journal of Coordination Chemistry
DOI: http://doi.org/10.1080/00958972.2017.1384547
Synthesis, characterization and DNA-intercalation studies of two ferrocene-
based Fe-Sn heterobimetallic compounds, and crystal structure of
trimethyltin (p-ferrocenyl)benzoate
ATAF ALI ALTAF* ¶†, NASIR KHAN¶ ‡, BHAJAN LAL§ and AMIN BADSHAH*‡
†Department of Chemistry, University of Gujrat, Hafiz Hayat Campus Gujrat - 50700 Pakistan
‡Department of Chemistry, Quaid-i-Azam University, Islamabad-45320, Pakistan
§Department of Energy System Engineering, Sukkur Institute of Business Administration, Sukkur, Pakistan
In this article, we report the synthesis of two (A3 and A4) new ferrocene-based heterobimetallic
compounds of Fe and Sn. Both compounds are structural isomers with formula
(C5H5)Fe(C5H4C6H4COO)Sn(CH3)3 and have been characterized by elemental analysis, cyclic
voltammetry, FTIR, UV-vis and NMR spectroscopy. Additionally, the structure of A3 was
confirmed by single-crystal XRD studies. XRD studies confirmed that in the solid phase A3
exists as a unidirectional polymeric material. DNA-binding potential of these compounds was
evaluated by UV-vis spectroscopy, cyclic voltammetry and viscosity measurements. These
interaction assays conclude that A3 (binding constant ~104 M-1) and A4 have good attraction
towards DNA and bind with DNA in intercalative manner.
Keywords: Heterobimetallic; DNA Interaction; Ferrocene; X-ray crystallography; Iron-tin
bimetallic
INTRODUCTION
The foremost reason for the numerous fatal diseases, for instance cancer, diabetes, hemophilia,
etc., is directly related to unrestrained production of proteins or transformed (misfolded or post-
translationally modified) proteins. DNA is a coding matter for proteins and its interactions with
*Corresponding authors. Email: aminbadshah@yahoo.com (A. Badshah); atafali_altaf@yahoo.com (A.A. Altaf)
¶These two authors contributed equally.
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drugs can affect the replication processes and serve as potential treatments for such diseases.
Even though before the discovery of DNA structure it was the target for cancer drugs [1, 2].
Compounds with prospective anticancer activity revealed later to target DNA either directly or
through inhibition of enzymes that control DNA integrity or provide building blocks for DNA.
There were several established therapeutic modalities targeting DNA; antimetabolites, these are
known to trim the number of nucleotides [3]; alkylating agents, those cause direct DNA-damage
and intercalators such as actinomycins, which bind to DNA and inhibit the activity of many
enzymes that use DNA as a substrate. Among the most widely and successfully used anticancer
agents today there are nonspecific DNA-targeting chemicals [4].
The use of metal-based compounds as anticancer agents represents one of the most
successful applications of bioinorganic chemistry. The DNA-binding properties of such
compounds play a vital role in their anticancer activity [5, 6]. The chemistry of organotin
compounds is very versatile in reference to their utility for cancer treatment and to target DNA
[7, 8]. Large numbers of reports are available in this context [9-11]; however, the ferrocene-
based heterobimetallics of tin are very limited [12]. Ferrocene plays an important role in the
development of anticancer drugs [13, 14]. For example, well-known anticancer agents, like
tamoxifen, are considered to be replaced by ferrocene analog ferrocifen [15]. Therefore, we
considered the mixing of ferrocene with organotin compounds in the form of heterobimetallic
molecules and subjected these molecules for interaction with DNA to evaluate their potential as
anticancer agents.
EXPERIMENTAL
Materials
Ferrocene, trimethyltin chloride, hexadecyltrimethylammonium bromide, sodium nitrite,
hydrochloric acid, 3-aminobenzoic acid and 4-aminobenzoic acid were acquired from Sigma-
Aldrich and used without purification. Solvents like acetone, chloroform, dichloromethane,
tetrahydrofuran, toluene, hexane, petroleum ether, ether and triethylamine were first distilled and
purified [16] and stocked up over molecular sieves of 4 Å.
Synthesis of 4-ferrocenylbenzoic acid (A1). A1 was synthesized according to literature reported
method [17, 18]; here we have given the protocol for the ease of the readers. A solution of 4-
2

aminobenzoic acid (7.0 g, 50 mmol) and 12 mL of HCl was prepared in 80 mL distilled water.
At 0-5 °C, 20 mL aqueous solution of sodium nitrite (3.5 g, 50 mmol) was added dropwise. After
complete addition, this diazonium salt solution was further stirred for 30 min by keeping the
temperature below 5 °C. A solution of ferrocene (10 g, 50 mmol) and 0.5 g
hexadecyltrimethylammonium bromide in 100 mL diethylether was added, to the diazonium salt
solution dropwise by keeping the temperature at 0-5 °C. After addition, the resulting mixture was
further stirred at room temperature for 8 h. On completion of the reaction, solvent was
evaporated under reduced pressure. The thus obtained red solid was added to the 1% aqueous
solution of NaOH and heated to 90 °C; hot solution was filtered to remove the excess of
ferrocene. The filtrate was allowed to cool to room temperature and equal volume of 1% aqueous
solution of HCl was added. After cooling the orange-red precipitate (ppt) of 4-ferrocenylbenzoic
acid starts to settle down at the bottom of beaker. The ppt was dried to get the 4-ferrocenyl
benzoic acid (A1) as orange red solid. Yield: 64%; Elemental analysis: Cal. (%) for C17H14Fe;
C, 66.70; H, 4.61; Found: C, 66.58; H, 4.73. FT-IR (υ, cm-1): 3500-2900, 2930, 2878, 1699
(CO2)sym, 1602, 1391(CO2)asym, 1106, 993, 853, 473 (Fe-Cp); 1H NMR (300 MHz, DMSO-d6)
(δ, ppm): 12.80 (s, 1H, COOH), 7.85 (d, 2H, J = 8.4 Hz, C6H4), 7.64 (d, 2H, J = 8.4 Hz, C6H4),
4.88 (t, 2 H, J = 1.8 Hz C5H4), 4.43 (t, 2H, J = 1.8 Hz, C5H4), 4.02 (s, 5H, C5H5); 13C NMR
(75 MHz, DMSO-d6) (δ, ppm): 167.7, 145.0, 129.9, 128.2, 126.0, 83.3, 70.1, 70.0, 67.2.
3-Ferrocenylbenzoic acid (A2). A2 was synthesized in the same way as described for A1; 3-
aminobenzoic acid was used instead of 4-aminobenzoic acid. Yield: 52%; Elemental analysis:
Cal. (%) for C17H14O2Fe; C, 66.70 H, 4.61; Found: C, 66.61; H, 4.51; FT-IR (υ, cm-1): 3442,
3014, 1681, 1605, 1468, 1414, 1331.5, 1260.9, 1101.4, 813.2, 479.4; 1H NMR (300 MHz,
DMSO-d6) (δ, ppm): 12.87 (s, 1H, COOH), 8.25 (s, 1H, C6H5), 7.98 (d, 1H, J = 7.8 Hz, C6H5),
7.74 (d, 1H, J = 7.8 Hz, C6H5), 7.43 (t, 1H, J = 7.8 Hz, C6H5), 4.74 (t, 2 H, J = 1.8 Hz, C5H4),
4.39 (t, 2 H, J = 1.8 Hz, C5H4), 4.08 (s, 5H, C5H5). 13C NMR (75 MHz, DMSO-d6) (δ, ppm):
171.2, 137.1, 133.5, 131.2, 130.0, 127.9, 126.2, 85.5, 70.0, 69.3, 68.6.
Trimethylstannyl 4-ferrocenylbenzoate (A3). Trimethyltin chloride (1.9 g, 10 mmol) was added
to a 250-mL round bottom flask charged with 4-ferrocenylbenzoic acid (3.1 g, 10 mmol) and
triethylamine (1.4 mL, 10 mmol) in 50 mL anhydrous toluene with constant stirring. The
3

reaction mixture was refluxed for 8 h and the extent of the reaction was monitored periodically
with TLC. After the completion of the reaction, the mixture was cooled to room temperature,
filtered and the filtrate was vacuum evaporated. Product A3 was re-crystallized from methanol as
orange red crystals. Yield: 70%; Elemental analysis: Cal. (%) for C20H22FeSnO2: C, 51.22; H,
4.73; Found: C, 51.34; H, 4.69; FTIR (υ, cm-1) 3091, 2927, 1602.4, 1582.2 (CO2sym), 1372
(CO2asym), 481.2 (Fe-Cp), 463.9 (Sn-C), 427.1 (Sn-O); 1H NMR (400 MHz, CDCl3) (δ, ppm):
7.99 (d, 2H, J = 8.4 Hz, C6H5), 7.52 (d, 2H, J = 8.4 Hz, C6H5), 4.63 (t, 2H, J = 1.8 Hz, C5H4),
4.30 (t, 2H, J = 1.8 Hz, C5H4), 4.08 (s, 5H, C5H5), 0.67 (s, 9H, -CH3), (SnMe), 2J [117/119Sn–1H]
[56/58 Hz]; 13C NMR (100 MHz, CDCl3) (δ, ppm): 171.7, 144.1, 130.3, 128.9, 125.4, 83.6, 69.8,
69.6, 66.8, -2.1; 1J[119/117Sn-13C] [398/380].
Trimethyltin(IV) 3-ferrocenylbenzoate (A4). A4 was synthesized in the same way as described
for A3; 3-ferrocenylbenzoic acid (A2) was used instead of 4-ferrocenylbenzoic acid (A1). Yield:
61%; Elemental analysis: Cal. (%) for C20H22FeSnO2; C, 51.22; H, 4.73, Fe, 11.91, Sn, 25.31;
Found C, 50.98; H, 4.80, Fe, 11.89, Sn, 24.99; FTIR (υ, cm-1) 3041, 2957, 1661 (CO2sym), 1602,
1402 (CO2asym), 481 (Fe-Cp), 463.9 (Sn-C), 422.1(Sn-O); 1H NMR (300 MHz, CDCl3) (δ, ppm):
8.16 (s, 1H, C6H5), 7.89 (d, 1H, J = 7.8 Hz, C6H5), 7.65 (d, 1H, J = 7.8 Hz, C6H5), 7.35 (t, 1H, J
= 7.8 Hz, C6H5), 4.72 (t, 2H, J = 1.8 Hz, C5H4), 4.35 (t, 2H, J = 1.8 Hz, C5H4), 4.06 (s, 5H,
C5H5), 0.68 (s, 9H, -CH3), 2J[119/117Sn-1H] [60]; 13C NMR (75 MHz, CDCl3) (δ, ppm): 171.8,
139.5, 131.8, 130.9, 129.7, 128.5, 127.7, 84.4, 69.6, 69.1, 66.5, -2.2. 1J[119/117Sn-13C] [398/380].
Instrumentation
1H and 13C NMR spectra were collected on Bruker 300 MHz / 400 MHz spectrometer with
deuterated solvents as an internal reference. Internal standard solvent; DMSO-d6: 1H = 2.50 and
13C = 40.11 ppm, CDCl3-d1: 1H = 7.28 and 13C = 77.05 ppm from TMS (Si(CH3)4). The
resonance splitting pattern in the 1H NMR spectra are defined as s = singlet, d = doublet, t =
triplet, and m = multiplet, etc., while coupling constants are reported in Hz. Elemental analysis
was carried out with a CHNS/O Analyzer (Thermo scientific Flash 2000 Organic elemental
analyzer) and metal contents were determined by Fissions Horizon Inductively Coupled Plasma
Optical emission Spectroscopy (ICP-OES). IR spectra were recorded by a FT-IR Bio-RAD
Merlin Varian / Thermo Scientific Nicolet-6700 FTIR spectrometer (400-4000 cm-1). Cyclic
4

voltammetry (CV) measurements were performed by an Eco Chemie Auto lab PGSTAT 12
potentiostat / galvanostat (Utrecht, The Netherlands) with the electrochemical software package
GPES 4.9.
X-ray single-crystal analysis: X-ray measurements were made on a Bruker kappa APEXII CCD
diffractometer equipped with graphite monochromated Mo-Kα (λ = 0.71073 Å) radiation. For
data collection ω scans, a multi-scan absorption correction was applied. The structure was solved
using SHELXS97 (Sheldrick, 273 2008) [19]. The structure was refined by full 276 matrix least-
squares techniques using SHELXL-97 [20].
DNA-binding studies
Sodium salt of calf-thymus DNA purchased from Acros Organics was used. The stock solution
of DNA (200 µM) was prepared by using doubly distilled water and stored at 4 °C for one day
before use. The concentration of DNA was determined by UV absorbance at 260 nm (molar
absorptivity coefficient (ε) of DNA was taken as 6600 M-1 cm-1) [21, 22]. The stock solutions of
test compounds were prepared in 80% dimethylsulfoxide (i.e. 20% H2O: 80% DMSO) and all
samples were buffered at pH 6 with phosphate buffer (0.1 M NaH2PO4 + 0.1 M NaOH).
UV-vis spectrometry: Absorption spectra were measured on a UV-vis spectrometer Shimadzu
1800. The electronic spectrum of a known concentration of the test compounds (A3 and A4) was
obtained without DNA. The spectroscopic response of the same amount of the test sample was
then monitored by addition of various concentrations of DNA in the solution. All samples were
allowed to equilibrate for 10 min prior to every spectroscopic measurement [23, 24].
Cyclic voltammetric studies: Electrochemical measurements were performed in an Eco Chemie
Auto lab PGSTAT 12 potentiostat / galvanostat (Utrecht, The Netherlands) single compartment
cell with three electrodes and equipped with electrochemical software package GPES 4.9. The
electrodes system comprised of a glassy carbon working electrode (surface area of 0.071 cm2),
Ag/AgCl as the reference electrode and a counter electrode of platinum wire of thickness 0.5
mm. The voltammogram of the test compound solution (1 mM) was recorded after purging with
argon gas for 10 min prior to measurements. The experiments were then repeated for the constant
5

concentration of test sample while varying calf-thymus DNA (20 µM to 60 µM) concentration.
After each measurement the working electrode surface was cleaned. Tetrabutylammonium
perchlorate (TBAP) was used as supporting electrolyte [25-27].
Viscometry: The DNA interaction by viscosity measurements were performed using an Ostwald
viscometer at 25±1 °C. The time for flow was measured using a digital stopwatch in triplicate.
Data were presented as relative viscosity, (η/ηo)1/3 versus compound concentration (μmol), where
η and ηo are the viscosity of DNA solution in the presence and absence of test compound,
respectively. Viscosity values were calculated using equation η = (t − to)/to, where t and to are the
observed flow time of test solution and solvent (DMSO:H2O, 8:2), respectively [25].
RESULTS AND DISCUSSION
Ferrocenylbenzoic acids (A1 and A2) were prepared by treating the diazonium salt of respective
aminobenzoic acids with ferrocene in the presence of phase transfer catalyst adopting the similar
procedure given in literature [17, 18, 28]. The scheme of reactions has been presented in
scheme 1. The synthesized acids (A1 and A2) reacted with trimethyltin chloride in toluene and
triethylamine was used to capture hydrochloric acid produced during reaction. As a result of this
reaction, the iron-tin heterobiorganometallic compounds (A3 and A4) were made (scheme 1). All
the synthetic products (A1-A4) were well-characterized using various techniques like elemental
analysis, FTIR spectroscopy, 1H and 13C NMR spectroscopy. Additionally, A3 was also
characterized by single-crystal X-ray diffraction analysis. The elemental analysis results indicate
that all compounds have good correlation with their theoretical compositions and have high
grades of purity in the solid state.
6

Scheme 1. Synthetic scheme for trimethyltin(IV) ferrocenylbenzoates (A3 and A4). Phenyl
substituents are on para position in A1 and A3, whereas on the meta-position in A2 and A4.
Hexadecyltrimethylammonium bromide was used as PTC.
In the solution phase, the compounds A1-A4 were studied by multinuclear NMR spectroscopy,
whereas bimetallic compounds A3 and A4 were additionally studied by UV-vis spectroscopy and
cyclic voltammetry. The multinuclear NMR (1H and 13C) spectra were recorded in DMSO-d6 (for
A1 and A2) and CDCl3 (for A3 and A4), and the resultant data have been reported in the
Experimental section. The proton NMR spectra of compounds A1 and A2 portray the
substitution of benzoic acid at one of the Cp rings of ferrocene. Since single signal of ferrocene
segregates into three peaks following the substitution [29], the singlet emerges for unsubstituted
C5H5 of ferrocene at 4.02-4.10 ppm corresponding to five protons. Whereas the derivatized Cp
signal separates out into two signals at 4.43 and 4.88 ppm for A1, and at 4.39 and 4.74 ppm for
A2. Together these bands exhibit triplet multiplicity with a coupling constant (J) value of 1.8 Hz,
which is evident of coupling owing to chemical equivalent protons. While the aromatic protons
from the phenyl ring of the acid resonate in their normal region. All the specific protons of these
complexes are identified by the amount and diversity of the spectral pattern which aids in
confirming the constitution of the present molecules under study. The 13C NMR spectra for A1
and A2 illustrate the substitution of benzoic acid moiety at ferrocene’s cp ring with one of the
hydrogens, as the single signal (in 13C NMR) originating from ferrocene is partitioned into four
peaks typical of mono-substituted ferrocene [29].
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The 1H and 13C NMR data for compounds A3 and A4 were taken in CDCl3 and reported
in the Experimental section with other data. Figure 1 presents the 1H NMR spectrum of A4 in
which trimethyltin signal appeared at 0.69 ppm with satellite signals for Sn–H J2-coupling of
58/56 Hz for 119/117Sn. This value of coupling constant indicates the distorted tetrahedral
geometrical arrangement around the tin center in the solution phase [30, 31]. The signals in the
region of 4.0-4.8 ppm and 7.3-8.2 ppm are characteristic of ferrocenyl and phenyl protons,
respectively, as in full agreement with literature data [17, 32-34]. The data recorded for A3 are
also of the similar pattern and justify its expected structure in the solution phase.
Figure 2 presents the 13C NMR spectrum of A4 in which trimethyltin signal appeared at
-2.2 ppm with satellite signals for Sn–C J1-coupling of 398/380 Hz for 119/117Sn. This value of
coupling constant also confirms the distorted tetrahedral geometrical arrangement around the tin
center in the solution phase [30, 31]. The signals in the region of 66.6-84.5 ppm and 127.5-
139.5 ppm are characteristic of ferrocenyl and phenyl group carbons, respectively, as in full
agreement with literature data [34-37]. The solvent signal appeared at 77.0 ppm whereas the
most downfield carbon of carboxylate group appeared at 171.8 ppm [38, 39]. The data recorded
for A3 are also of the similar pattern and justify its expected structure and stability in solution
phase in accordance with other characterization techniques.
The UV-vis absorption spectra of 40 µM A3 and 40 µM A4 was scanned in the region
220-700 nm. The UV-vis spectrum of A3 is presented in figure 3 (gray-colored line) and
characterized with four signals at 256, 281, 350 and 450 nm. The molar absorption coefficients
for these signals are 20750, 21750, 4825 and 2125 M-1 cm-1, respectively. The absorption band at
254 nm may arise from n-π* transition of -COOH owing to electron-donating character of
ferrocenyl group, while the 296-nm band is related to π-π* para-disubstituted phenyl group. The
low molar absorptivity bands at 350 and 454 nm are attributed to charge-transfer and d-d
transitions in the ferrocene part, respectively [38, 40]. The UV-vis spectrum of A4 is presented in
figure 4 (skyblue-colored line) and characterized with three signals at 280, 340 and 430 nm. The
molar absorption coefficients for these signals are 24750, 4100 and 2050 M-1 cm-1, respectively.
The absorption band at 296 nm is related to π-π* meta-disubstituted phenyl group. The low
molar absorptivity bands at 340 and 430 nm are attributed to charge-transfer and d-d transitions
in the ferrocene part, respectively [38, 40].
8

To understand the electrochemical behavior of synthesized compounds A3 and A4 in the
solution, these compounds were characterized by cyclic voltammetry on a three electrode
system. The voltammogram of A3 (1 mM in 20% aqueous DMSO buffer at pH 6.0; supporting
electrolyte 0.1 M TBAP) depicts distinct and stable redox peaks within 0.0 to 0.60 V potential
ranges with a quasi-reversible system (shown in figure 5, green line), with electrode potential
difference (ΔEp = Epc - Epa) of more than 57 mV and (Ipa/Ipc) current ratio is not equal to one,
where Epc, Epa, Ipc and Ipa are cathodic peak potential, anodic peak potential, cathodic peak
current and anodic peak currents, respectively. These oxidation (Epa = 0.28 V) and reduction
signals (Epa = 0.21 V) can be attributed to the ferrocene and ferrocenium reversible change, as
reported in the literature. In literature, for the ferrocene in its derivatives, the redox couples
normally generate the signal in the range of 0.20 to 0.70 V [41-43], whereas the pure ferrocene
generate oxidation signal at 0.52 V and reduction signal at 0.46 V under similar conditions
[43-45]. The slight shift to the negative side in the redox potential of A3 can simply be
understood on the basis of the electron-donating group’s attachment in the structure. In case of
A4 (shown in figure 6, red line), these oxidation (Epa = 0.55 V) and reduction signals (Epa =
0.25 V) shifted less in comparison to simple ferrocene. This is again understandable as
substituent at meta-position always effect less, to the electronic charge densities, than the para-
ones [42].
Solid phase FT IR spectroscopy along with elemental analysis was utilized for
characterization, and A3 was additionally studied by single-crystal X-ray diffraction analysis.
Compounds having covalent bonding with induced net dipole moment absorb variant infrared
electromagnetic radiations from 4000-400 cm-1. The synthesized compounds were analyzed by
FT-IR and the spectra values of the complexes were compared with literature values of
associated compounds. The significant/specific bands for the synthesized compounds are given
with other characterization data in the Experimental section. For A1 and A2, emergence of a
single broad peak centered around 3300 cm-1 provide information for the existence of OH group
of ferrocenylbenzoic acids, while sp3 and sp2 carbon protons of the aromatic and ferrocene
groups appear in their specific vibration regions. The IR spectra of the synthesized organotin
ferrocenylbenzoates have been recorded. The vibrational spectral data provide evidence for the
molecular constitution of A3 and A4. In all compounds, the disappearance of ν(O-H) in the range
of 3300 cm-1 supports the deprotonation of ferrocenylbenzoic acid and the shift of signals in the
9

functional group region, i.e. 1680-1350 cm-1 due to carbonyl carbon, are owing to the interaction
between the carbonyl oxygen atoms of the carboxylate groups and the tin atom [46]. In the IR
spectra of A3 and A4, the stretching frequencies of Sn-O formation appear between 450-
415 cm-1, Fe-cp appear in the region of 500-460 cm-1, while Sn-C absorption frequencies are
observed between 490-443 cm-1, consistent with the literature values [47-49].
Single-crystal X-ray diffraction analysis of A3: An X-ray diffraction study of A3 was carried out
(relevant details can be found in the Instrumentation section). A3 was crystallized as orange
block-like crystals from CHCl3:C2H5OH (1:1) mixture. The crystal system and space group of
A3 was found as monoclinic and P21/c, respectively. The other crystal parameters found, at
100 K using Mo K (λ = 0.71073) radiations, were: empirical formula C20H22FeO2Sn, unit cell
dimensions a = 6.4454(5) Å, b = 9.6305(8) Å, c = 29.030(2) Å,  = 92.169(4)°, Mr = 468.92,
volume = 1800.7(3) Å3, Z = 4, density = 1.513 g/cm3, F(000) = 1048.0, Crystal size = 0.22 ×
0.18 × 0.03 mm3, Index ranges (h, k, l)max = (8, 0, 38), (h, k, l)min = (0, -12, -38), Total reflections
= 5176, Mu = 2.21 mm-1, θmax = 28.27°, R = 0.0471 and wR2 = 0.1058, S = 1.284.
Figure 7 presents the ORTEP diagram for the monomer unit of A3 with numbering
scheme and table 1 contains selected important bond lengths, bond angles and torsion angles.
Compound A3 is crystallized in the polymeric structure as shown in figure 8. The polymeric
nature and trans-O2SnC3 coordination sphere is common for R3SnO2CR` (having R = n-alkyl
groups) [50, 51]. The nature of the bridging is anisotropic, in which tin forms one short
[Sn(1)-O(1) = 216.1(4) pm] and one relatively long [Sn(1)-O(2) = 244.3(4) pm] bond to oxygen.
These bond lengths are comparable to other Sn-O bonds found in typical polymeric triorganotin
carboxylates, with short (212.0-224.6 pm) and long (224-265 pm) bonds, respectively. The
O(1)-Sn(1)-O(2) angle [= 172.69(13)] is also comparable to other angles found in typical
polymeric triorganotin carboxylates (168.6-178.7) [51]. Ferrocene, on the one dimensional zig-
zag polymeric structure, directed upward and downward alternative to the tin carboxylate plane.
This setup mediates a metal-organic network as shown in figure 8.
It has been stated that there is a clear preference for the polymer structure of compound
A3 type, except in the case of excessive bulk, either at the tin-bound organic group or at the
carboxylate moiety, in which case the other type is favored as reported in the literature [52].
10

While it is generally true that bulky organic groups at tin preclude the adoption of the polymeric
motif [53, 54].
DNA-binding studies
Binding of small molecules to DNA can be detected by a number of techniques. The present
report discusses DNA-binding studies of new compounds using various techniques like UV-Vis
absorption spectroscopy, cyclic voltammetry and viscometry. To avoid the decomposition of the
ferrocenium state if formed in basic or even neutral solution and protonation of the ferrocenyl
group in acidic conditions, these studies have been carried out in phosphate buffer at pH = 6.8
[45].
UV-vis spectroscopic titration: The binding empathy of ferrocene containing organotin
derivatives A3 and A4 with DNA was evaluated by spectral variations on UV-vis spectrum. This
is an important technique to evaluate interaction of small molecules with DNA. As a result of
interaction between small molecules and DNA, there always occur changes in the energy of
HOMO and LUMO levels of DNA and the small molecules. UV-vis spectroscopy is useful in
probing such energy changes [55]. The UV-vis absorption spectra of 40 µM A3 in the absence
and presence of DNA (various concentrations) is presented in figure 3.
Compound A3 is marked with four signals as described in previous sections. Changes in
the electronic spectrum of A3 with continuous addition of DNA were observed. After the
addition of DNA, there is increase in absorbance for signal at 254 nm and decrease in absorbance
for signals at 350 and 450 nm. The increase about 254 nm can be attributed to the increasing
concentration of DNA, as DNA nucleotides also absorb in the same region. However the
decrease in absorbance about 350 and 450 nm signals is important to understand the interaction
between A3 and DNA. Some shifts (about 8 to 10 nm towards lower energy) in the λmax for these
signals were also observed. Such red-shift in the λmax and hypsochromism for signals about
350 nm and 450 nm indicate the intercalation of the planar parts of A3 between DNA-base pairs,
as similar changes were always observed in the electronic spectrum of intercalators in literature
[56, 57].
Figure 4 presents the changes in the electronic spectrum of A4 upon interaction with
DNA. There are no considerable changes in the electronic spectrum of A4 after the addition of
11

DNA, except the increase in absorbance of the signal about 250 nm. This increase in absorbance
is mainly due to the intrinsic absorbance of DNA in that region. These results indicate poor or no
interaction between A4 and DNA; it might be due to the non-planar structure of A4. UV-vis
spectroscopy is also an effective tool for quantification of binding strength of DNA with metal
complexes [58, 59]. Following host guest equation 1 can be utilized for this purpose:
Ao/(A-Ao) = εF / (εB – εF) + εF / (εB – εF)*1/K [DNA]
(1)
where K is the binding constant, Ao and A is the absorbance of the free compound and the
apparent one after addition of DNA, εF and εB are their absorption coefficients, respectively. The
slope to intercept ratio of the plot between Ao/(A−Ao) versus 1/[DNA] yielded the binding
constant K which is reported in table 2 and compared with the other literature reported similar
compounds. Using the value of K for the measure at room temperature (T = 25 C), the value of
change in Gibbs free energy (ΔG) was calculated using the equation ΔG = -RT lnK. The high
negative value of ΔG indicates the spontaneity of the reaction between tested compounds and
DNA [60].
Scheme 2. Structures for the example compounds (Ex-1 to Ex-3) taken from literature for the
comparison of DNA binding activities.
Cyclic voltammetry: Cyclic voltammetry measurements were executed with the objective to
consider the redox behavior of electroactive compounds with DNA and their binding affinity
12

using a three electrode system [63]. The voltammograms of complexes depict distinct and stable
redox peaks within 0.0-0.7 V potential range with a quasi-reversible system, as discussed in
previous section. The CV measurements were conducted at a steady concentration of compounds
(1 mM) and rising the concentrations DNA (40-60 µM). The incremental addition of DNA to a
1 mM solution of complex A3 is accompanied by a change in peak potential and a decrease in
peak current (ipa and ipc). The current reduction is owed to the amalgamation of the complex and
DNA producing supramolecular species which results in lowering of diffusion of electro-active
species towards electrode, hence decreasing the electrolytic current [43].
The shift in peak potential is used to investigate the mode of interaction between small
molecules and DNA [64]. The negative shift in the peak potential is indicative of the presence of
electrostatic interaction of the cationic drug with the anionic phosphate of DNA backbone
whereas a positive shift in the peak potential is characteristic for the intercalation of drug into
double-helical structure of DNA [23]. Therefore, the positive shift in the potential for A3 is
attributed to the intercalation of the molecule into the double-helix of DNA. As it is evident that
the size of ferrocene restricts its penetration into the DNA, so it is plausible that other groups
(most probably the phenyl group) intercalate into DNA. Molecule A4 also behaves in similar
pattern in cyclic voltammetry titration as shown in figure 6, but in this case the depression of
peak current and the change in peak potential is lower than that in the case of A3.
This behavior shows a similar interaction between DNA and each of A3 and A4; it
further provides evidence that the interaction of A4 is poorer than that of A3, which is in
agreement with the results obtained in case of UV-vis spectroscopy. In the case of A3, ferrocene
unit is bonded to the more planar spacer phenyl ring as it is evident from single-crystal data that
will increase its ability of intercalate with DNA. While in A4, the presence of two groups on
meta-position restrict completely the penetration of the complexes and other groups into the
double-helix of DNA resulting in poor interaction. The drop in current is attributed to the slow
diffusion of supramolecular complex formed between the compound and DNA. The binding
constant (K) is determined using the following equation (2) [65]:
13

where i and io are the peak currents for the compound in the presence and absence of DNA,
respectively, and A is the proportionality constant. The binding constant K was calculated from
the intercept of the plot 1/[DNA] versus 1/(1-i/io). The value of K calculated in this way has been
reported in table 2. The formal potential difference for the free and bonded compound can be
used to find either ferrocene or ferrocenium form is interacting. In the case of A3, the reduced
form is more active as evident from figure 5 and the value of Kred/Koxd (Kred binding constant for
reduced species and Koxd binding constant for oxidized species) calculated from equation (3)
[36, 45]:
(3)
where Eb° and Ef° are the formal potentials of the free and bound forms of small molecule,
respectively. The positive shift indicates that the reduced form interacts strongly with DNA for
A3 and A4. So, the interaction of DNA probably entail that Fe2+ first combines with DNA
followed by the generation of Fe3+ and production of reactive oxygenated species (ROS)
resulting in DNA-damage oxidative stress as elaborated in scheme 3 [66].
Scheme 3. The general redox process of the free and DNA-bound compounds, the experimental
results show the lower equilibrium is more favorable in current study.
Viscometry: To further confirm the DNA binding mode, viscometry experiments were performed
for both compounds. The results obtained in these measurements are shown in figure 9.
Hydrodynamic measurements are the important test for the understanding the binding mode in
small molecule-DNA interaction. Simply the increase in viscosity of the DNA solution suggests
14

the intercalation of small molecule into the DNA helical structure whereas a decrease in viscosity
is interpreted to the electrostatic interaction [26, 62].
As shown in figure 9, there is notable increase in the relative viscosity of DNA upon
increasing concentration of A3. However in the case of A4, there is relatively less or no increase
in viscosity. The increase in relative viscosity indicates the intercalative mode of binding for the
test compounds. So, these results also support the observation of UV-vis spectrum and the
electrochemistry of the reaction. Such interaction can cause conformational changes in DNA
structure, which escorts to the dysfunction of normal DNA and may prevent its replication and
lead to cell death [66].
CONCLUSION
Two new heterobimetallics (of Fe and Sn) A3 and A4 have been synthesized and characterized.
J-values (2J [117/119Sn–1H] [56/58 Hz]) for tin–proton coupling indicate the distorted tetrahedral
geometrical arrangement around the tin center in the solution. XRD studies confirmed that in the
solid phase, they exist as unidirectional polymeric materials. That characteristic data ensures the
stability of formed complexes in solid as well as in the solution. DNA-binding potential of these
compounds was evaluated by different physicochemical methods. These interaction assays
conclude that A3 and A4 have good affinity towards DNA and bind with DNA in intercalative
manner. The binding constant of the compound with 3-ferrocenylbenzoate (A4) is about 10 times
lower than the corresponding 4-ferrocenylbenzoate (A3). The comparison of results with
reported similar compounds (shown in scheme 2) indicate that A3 has highest DNA-intercalation
ability among the heterobimetallic compounds; this activity may be attributed to linearity and
less crowded structure of A3. Such interaction may induce perturbation in the normal functioning
of DNA which could presumably culminate in the prevention of replication and ultimate cell
death. Therefore, these compounds can be potential anticancer agents in the future.
Supplementary material
The crystallographic data for A3 (CCDC No. 1562612) can be obtained from the Cambridge
Crystallographic Data Centre via free of charge from www.ccdc.cam.ac.uk and the CIF file for
the same has also been provided in the supplementary data.
15

Acknowledgement
The authors are thankful to the Higher Education Commission (HEC) Pakistan for the financial
support.
Competing interests
The authors have no competing interests.
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19

Figure 1. 1H NMR spectrum of A4 in CDCl3 referenced with internal standard TMS.
20

Figure 2. 13C NMR spectrum of A4 in CDCl3 referenced with internal standard TMS.
21

Figure 3. Electronic absorption spectra of 40 µM compound A3 without DNA and with 22.5-
80 µM DNA in 20% H2O: 80% DMSO buffered at pH 6.0.
22

Figure 4. DNA-interaction electronic absorption spectra of compound A4 (40 µM) without DNA
and with 34–80 µM DNA in 20% H2O: 80% DMSO buffered at pH 6.0.
23

Figure 5. Cyclic voltammograms of A3 (1 mM at 298 K) recorded at 100 mV s-1 potential sweep
rate on glassy carbon electrode in the absence and presence of increasing concentration of DNA
(40 µM and 60 µM) in 20% H2O: 80% DMSO. All solutions were buffered at pH 6.0 and 0.1 M
TBAP was used as supporting electrolyte.
24

Figure 6. Cyclic voltammograms of 1 mM A4 recorded at 100 mV s-1 potential sweep rate on
glassy carbon electrode at 298 K in the absence and presence of 1 mL of increasing
concentration of CT-DNA (40 µM and 60 µM) in 20% H2O: 80% DMSO buffer at pH 6.0;
supporting electrolyte 0.1 M TBAP.
25

Figure 7. ORTEP diagram of A3 with numbering scheme (hydrogens are presented in cyan
color).
26

Figure 8. Polymeric structure of A3 in solid structure formed by O-Sn-O linkage and bear
ferrocene in zigzag manner.
27

Figure 9. Viscometric results of compounds A3 and A4.
28

Table 1. Selected bond lengths (Å), bond angles () and torsion angles () of A3.
Bond lengths (Å)
Bond angles ()
Torsion angles ()
C(1)-Sn(1)
C(4)-O(2)
C(4)-O(1)
C(8)-C(11)
C(11)-Fe(1) 2.053(5)
C(16)-Fe(1) 2.041(3)
2.110(4)
Sn(1)-C(1)-H(1)
O(2)-C(4)-O(1)
O(2)-C(4)-C(5)
O(1)-C(4)-C(5)
O(1)-Sn(1)-O(2)
C(1)-Sn(1)-O(1)
C(1)-Sn(1)-O(2)
C(12)-Fe(1)-C(20)
109
O(2)-C(4)-C(5)-C(10)-16.2(7)
O(1)-C(4)-C(5)-C(10)163.6(5)
1.244(6)
1.273(6)
1.480(7)
124.0(5)
120.8(4)
115.2(4)
172.69(13)
88.47(17)
84.49(17)
161.5(2)
C(7)-C(8)-C(11)-C(15)
34.9(7)
-
C(4)-C(5)-C(10)-C(9) -177.2(5)
O(2)-C(4)-O(1)-Sn(1) -5.0(7)
C(5)-C(4)-O(1)-Sn(1) 175.1(3)
O(1)-C(4)-O(2)-Sn(1) 125.9(6)
C(4)-O(1)-Sn(1)-O(2) 77.0(10)
Sn(1)-O(1)
Sn(1)-O(2)
2.161(4)
2.443(4)
29