Synthesis, characterization and biological evaluation of ruthenium flavanol complexes against breast cancer

Article abstract of DOI:10.1016/j.saa.2017.02.056

Four Ru(II) DMSO complexes (M1R-M4R) having substituted flavones viz. 3-Hydroxy-2-(4-methoxyphenyl)-4H-chromen-4-one (HL1), 3-Hydroxy-2-(4-nitrophenyl)-4H-chromen-4-one (HL2), 3-Hydroxy-2-(4-dimethylaminophenyl)-4H–chromen-4-one (HL3) and 3-Hydroxy-2-(4-chlorophenyl)-4H-chromen-4-one (HL4) were synthesized and characterized by elemental analysis, IR, UV–Vis, ¹H NMR spectroscopies and ESI-MS. The molecular structures of the complexes were investigated by integrated spectroscopic and computational techniques (DFT). Both ligands as well as their complexes were screened for anticancer activities against breast cancer cell lines MCF-7. Cytotoxicity was assayed by MTT [3-(4, 5-dimethyl thiazol-2-yl)-2, 5-diphenyl tetrazolium bromide] assay. All ligands and their complexes exhibited significant cytotoxic potential of 5–40?μM concentration at incubation period of 24?h. The cell cytotoxicity increased significantly in a concentration-dependent manner. In this series of compounds, HL2 (IC₅₀ 17.2?μM) and its complex M2R (IC₅₀ 16?μM) induced the highest cytotoxicity.

Full text of DOI:10.1016/j.saa.2017.02.056

Accepted Manuscript
Synthesis, characterization and biological evaluation of ruthenium
flavanol complexes against breast cancer
Ashok Kumar Singh, Gunjan Saxena, Sahabjada, M. Arshad
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DOI:
Reference:
S1386-1425(17)30167-1
doi: 10.1016/j.saa.2017.02.056
SAA 14978
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ACCEPTED MANUSCRIPT
Synthesis, Characterization and Biological evaluation of Ruthenium Flavanol
Complexes against breast cancer
Ashok Kumar Singha*, Gunjan Saxenaa , Sahabjadab, M. Arshadb
aDepartment of Chemistry, University of Lucknow, Lucknow – 226 007 INDIA
bDepartment of Zoology, University of Lucknow, Lucknow – 226 007 INDIA
*Corresponding author: Ashok Kumar Singh; Phone: +91-9919663856;
E-mail address: singhaks3@rediffmail.com
Abstract
Four Ru(II) DMSO complexes (M1R-M4R) having substituted flavones viz. 3-Hydroxy-
2-(4-methoxyphenyl)-4H-chromen-4-one (HL1), 3-Hydroxy-2-(4-nitrophenyl)-4H-chromen-4-
one (HL2), 3-Hydroxy-2-(4-dimethylaminophenyl)-4H-chromen-4-one (HL3) and 3-Hydroxy-2-
(4-chlorophenyl)-4H-chromen-4-one (HL4) were synthesized and characterized by elemental
1
analysis, IR, UV-Vis, H NMR spectroscopies and ESI-MS. The molecular structures of the
complexes were investigated by integrated spectroscopic and computational techniques (DFT).
Both ligands as well as their complexes were screened for anticancer activities against breast
cancer cell lines MCF-7. Cytotoxicity was assayed by MTT [3-(4, 5-dimethyl thiazol-2-yl)-2, 5-
diphenyl tetrazolium bromide] assay. All ligands and their complexes exhibited significant
cytotoxic potential of 5 – 40 µM concentration at incubation period of 24 h. The cell cytotoxicity
increased significantly in a concentration-dependent manner. In this series of compounds, HL2
(IC50 17.2 µM) and its complex M2R (IC50 16 µM) induced the highest cytotoxicity.
Keywords: Ru(II); DMSO; Flavones; Cytotoxicity; DFT
1

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1. Introduction
During the last three decades, platinum-based drugs have played a crucial role in cancer
chemotherapy [1]. The cis-[PtCl2(NH3)2] complex, known as cis-platin is the most widely used
drug in anticancer therapy [2,3]. In recent years, the antitumor properties of ruthenium(II)
compounds
are
gaining
considerable
importance
and
a
(chloroimidazole)
ruthenium(III) complex had entered in phase I clinical study [4a]. The first ruthenium complex
investigated for anticancer activity was fac-[RuCl3(NH3)3] [4b]. This compound was not pursued
much further because its poor solubility precludes it from adequate formulation as a drug [5].
The anticancer activity of ruthenium(II) complex, cis-[RuCl2(DMSO)4] (DMSO = dimethyl
sulfoxide), is well-known and had been compared with cis-platin [6a]. Ruthenium anticancer viz.
ruthenium DMSO and ofloxacin ruthenium bipyridyl complexes based drugs [6b-e] have no side
effects when compared with platinum based drugs which is having relatively more side effects
such as nephrotoxicity and neurotoxicity as a result its use has been limited [7]. The metallo-
drugs based on ruthenium are often identified as less toxic and capable of overcoming the
resistance induced by platinum drugs in cancer cells [8]. Due to these properties several
ruthenium based anticancer agents, ruthenium-DMSO complexes and the coordination
complexes of ruthenium-chloro-DMSO containing heterocyclic ligands viz. NAMI, have proved
their potential in the treatment of cancer cells [9]. The cis-[Ru(DMSO)4Cl2] has been widely
used as a precursor for the synthesis of Ru(II) complexes by displacement of DMSO molecules
with variety of ligands [10]. Ruthenium drugs are gaining importance in clinic due to their low
toxicity and specific activity than the current metallo-drugs [11]. Furthermore, ruthenium
complexes appear to penetrate into the tumor cells where they bind effectively with DNA [12].
The two ruthenium compounds, ImH [Ru(im)CI5] [13], NAMI-A (im = imidazole) and Ru (III)
compound indazolium trans-[tetrachlorobis(1H-indazole) ruthenate (III)] KP1019 [14] were the
first ruthenium-based anticancer drugs which entered into clinical trials [15]. Also, many other
compounds that include ruthenium center have been developed and tested [16] viz., trans-
(IndH)[Ru(ind)2Cl (Ind=indazole), mer-Ru(tpy)Cl3(terpy=2,2’,6’,2”-terpyridine [17, 18],
ImH[Ru(im)CI5]andImH[Ru(im)2CI4] were found to be active as cytostatic drugs [19, 20].
Flavones are the yellowish polyphenolic compounds belonging to class flavonoids which exist in
plants and are having secondary metabolites, 2-phenyl benzopyran functionality. They are
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potential synthetic targets as they display wide range of biological activities [21-23], including
antibacterial [24], antioxidant [25], anti-inflammatory [26-28], antiallergic [29-31] and
antiestrogenic [32] activities. Flavonoids are benzo-γ-pyrone derivatives consisting of phenolic
and pyrone rings, and are classified into, flavones, flavonols, flavanones, isoflavones and
anthocyanidins. Flavonols are the most abundant flavonoids in foods [33]. The flavones also
known as anthoxanthins are yellow pigments, which is responsible for colour of flavones show
antioxidant properties [34]. It may be possible that ruthenium and other transition metals can
form coordination complexes with flavones having different substituents at para- position.
Because of the differences in the electronic properties of these para-substituents the anticancer
activities of the resulting complexes against breast cancer may be tuned.
With these viewpoints and in the quest of new ruthenium(II)DMSO flavonol complexes having
potential against breast cancer cell lines herein we report the synthesis, characterization and
biological evaluation of four new ruthenium(II)DMSO complexes with substituted flavonols
against breast cancer.
2. Experimental
2.1 Materials and measurements
All chemicals were purchased from Aldrich-sigma, Himedia and E. Merck and used without
further purification. Solvents were purified and dried according to standard procedure [35]. IR
1
spectra were recorded on Perkin–Elmer AC-1 spectrometer and H NMR spectra were recorded
in DMSO-d6 on Bruker Avance IIIHD spectrometer at 400 MHz using TMS as an internal
reference. Elemental analysis were performed on Exeter analytical Inc.‘‘Model CE-440 CHNS
analyzer’’. ESI-MS spectrometry were performed on JEOL SX 102/DA-6000. Electronic
absorption spectra were recorded on LT 2900–UV-Vis spectrophotometer. Conductance
measurements were carried by using Esico model-1601. All biological activities were carried
out from Department of Zoology, University of Lucknow.
2.2 Cell culture
MCF-7 is an estrogen receptor positive breast cancer cell line derived from pleural effusion and
is the most commonly used cell line for screening of anticancer breast agents. MCF-7 cells
obtained from cell repository-NCCS Pune (Job No. 390) were cultured in Eagle's minimal
essential medium (MEM, Himedia) with NEAA, 1.5 g/L NaHCO3, 1 mM sodium pyruvate and
0.01 mg/mL insulin with 10% fetal calf serum (Himedia).
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2.3 Cell cytotoxicity (MTT assay)
The antiproliferative activities of the compounds were determined using MTT [3-(4, 5-
dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide, Himedia] assay according to Sharma et
al. [36]. In brief, 1×104 cells/well were seeded in 100 μL complete MEM (minimal essential
medium) in each well of 96-well culture plates and incubated for 24 h at 37oC in a CO2
incubator. Compounds were diluted to the desired concentrations in culture medium and added to
the wells with respective vehicle control. After 24 h of compounds supplementation, 10 μL of
MTT reagent (5 mg/mL) was added in each well and the plates were further incubated for 4 h at
37oC until purple formazan crystal produced. Supernatant was then removed from each well and
crystals were solubilized in 100 μL of DMSO reagent. The plate was left for 10 min at 37oC and
absorbance was then recorded at 540 nm by a microplate reader (BIORAD Model 680). For each
condition, at least three independent experiments were performed [36, 37]. The percentage
antiproliferative activities were calculated by using the formula-

OD of control  OD of treated

% Cell cytotoxici ty 
100

OD of control

The plot of % cell cytotoxicity versus compound concentration was applied to calculate the IC50.
2.4 Analysis of cellular DNA content and detection of apoptotic cells
MCF-7 cells were plated into 6-well plate at density 1×106 cells/mL and treated with a
concentration of IC50 value of highly active compounds HL1 and M1R for 24 h in 5% CO2
incubator and 37oC temperature. After 24 h incubation, the cultured cells were harvested and
washed with cold PBS and fixed in 70% ethanol, treated with RNAse A (10 mg/mL) and stained
with propidium iodide, PI (10 µg/mL) for 30 min at room temperature in dark. The PI
fluorescence of individual nuclei was measured using a FACS-Calibur cytometer (BD
Biosciences, Heidelberg, Germany). Data were analyzed with the Cell Quess Pro V5.2.1
software (BD Biosciences).
2.5 Statistical analysis
The results are presented in mean ± SE. The unpaired t-test was used to compare the differences
between parameters at different concentrations. The one way analysis of variance (ANOVA) was
used to compare the differences within parameters among concentrations. The Bonferroni pair
wise comparison test was applied to compare the differences between concentrations for a
parameter if found significant in ANOVA. The p-value <0.05 was considered to be as
significant.
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2.6 Synthesis: 3-Hydroxy-4-substituted flavonols HL1-HL4 and their metal complexes M1R-M4R
Step A: Chalcones were synthesized by Claisen-Schmidt reaction by previously reported methods.
The synthesis of compounds were carried out in dilute ethanolic sodium hydroxide solution by
cyclization and Claisen- Schmidt condensation of 2-hydroxyacetophenone with different 4-
substituted benzaldehyde [38,39,40] at room temperature. The equimolar ratio of 2-hydroxy
acetophenone (10 mmol, 1 mL) with 4-substituted benzaldehyde (10 mmol, 1 mL (for HL1)/ 1.52
g (for HL2)/ 1.50 g (for HL3)/ 1.41g (for HL4)) in 30 mL of ethanol were stirred and 40% NaOH
solution (10 mL) was added dropwise and the mixture was stirred for 4 days. After stirring for 4
days the solution was diluted with water, acidified with 10% HCl, and kept overnight. The crude
products were filtered, washed with diethyl ether and recrystallized in ethanol to get the desired
chalcones. Chalcones (1 mmol; 0.254 g for HL1)/ 0.269 g (for HL2)/ 0.267 g (for HL3)/ 0.258 g
(for HL4)) were dissolved in ethanol (15 mL) and stirred and 10% (15 mL) NaOH was added. To
the resulting mixtures 30% H2O2 (10 mL) was added and the precipitate was recovered by addition
of dichloromethane. The obtained flavonols were recrystallized using ethanol.
3-Hydroxy-2-(4-methoxyphenyl)-4H-chromen-4-one (HL1): Colour: Reddish Brown; Yield
1
(55%, 0.140 g); m. p. 135oC; IR (KBr/cm-1): 1684 νC=O, 3500 νOH, 1283 δOH, 1591 νC=C; H
NMR (400 MHz, DMSO-d6, 25oC, TMS): δ 10.67 (s, 1H, OH), δ 6.99-7.73( multiplet, Ar-H), δ
2.4 (s, 6H, OCH3); For C16H12O4; ESI/MS (268). Calcd: C 71.64, H 4.48, Found: C 72.03, H
4.72.
3-Hydroxy-2-(4-nitrophenyl)-4H-chromen-4-one (HL2): Colour: Dark Brown; Yield (60%,
1
0.169 g); m. p. 138oC; IR (KBr/cm-1): 1693 νC=O, 3366 νOH, 1235 δOH, 1520 νC=C; H NMR
(400 MHz, DMSO-d6, 25oC, TMS): δ 10.86 (s, 1H, OH), δ 7.16-7.50 (Ar-H); For C15H9NO5;
ESI/MS (283). Calcd: C 63.60, H 3.18, N 4.95 Found: C 63.10, H 3.32, N 4.68.
3-Hydroxy-2-(4-dimethylaminophenyl)-4H-chromen-4-one (HL3): Colour: Brown; Yield
1
(62%, 0.187 g); m. p. 140oC; IR (KBr/cm-1): 1692 νC=O, 3475 νOH, 1273 δOH, 1568 νC=C; H
NMR (400 MHz, DMSO-d6, 25oC, TMS): δ 13.35 (s, 1H, OH), δ 7.14-7.50 (Ar-H), δ 3.68 (s,
6H, CH3).; For C17H15NO3; ESI/MS (281). C 72.60, H 5.34, N 4.98 Found: C 72.96, H 5.52, N
4.74.
3-Hydroxy-2-(4-chlorophenyl)-4H-chromen-4-one (HL4): Colour: Yellow; Yield (65%, 0.154
g) ; m.p. 135oC; IR (KBr/cm-1): 1699 νC=O, 3423 νOH, 1209 δOH, 1566 νC=C; 1H NMR (400 MHz,
5

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DMSO-d6, 25oC, TMS): δ 10.57 (s, 1H, OH), δ 7.22-7.73 (Ar-H); For C15H9O3Cl; ESI/MS (272).
C 66.18, H 3.31, Found: C 65.83, H 3.62.
Step B: [Ru(DMSO)4Cl2] was prepared according to the method reported by Evans [10]. The
synthesis of [Ru(DMSO)4Cl2] was carried out by mixing of RuCl3.3H2O and 1 mL of
dimethylsulphoxide and heating the mixture for 1 h. The yellow crystals of [Ru(DMSO)4Cl2]
was washed with ether then was used as such. All the complexes were prepared under nitrogen
atmosphere. The mononuclear complexes were synthesized by drop-wise addition of ethanol
solution of ligands HL1-HL4 (1 mmol; 0.268 g (HL1)/ 0.283 g (HL2)/ 0.281 g (HL3)/ 0.272 g
(HL4)) to the ethanolic solution of [Ru(DMSO)4Cl2] (1 mmol; 0.484 g). After complete addition
of ligand one drop of triethylamine was added to the resultant mixture and whole mixture was
refluxed with stirring for 7 h. The completion of reaction was monitored through TLC. After 7 h
the contents were cooled to room temperature and filtered. To the filtrate a saturated methanolic
solution of sodium nitrate was added. The brown precipitate thus obtained was filtered and
washed with water, methanol followed by ethanol/ diethyl ether and then purified by column
chromatography over alumina using CH3CN as eluent. The molecular formula of the complexes
are [Ru(DMSO)2(L1)2] 2NaNO3.2H2O (M1R); [Ru(DMSO)2(L2)2].2NaNO3.H2O (M2R);
[Ru(DMSO)2(HL3)2](NO3)2.2H2O (M3R); and [Ru(DMSO)2(L4)2].2NaNO3.5H2O (M4R). The
conductivity measurements of metal complexes show that M1R, M2R and M4R are non-
electrolytic but M3R is electrolytic in nature.
[Ru(DMSO)2(L1)2] 2NaNO3.2H2O (M1R): Colour: Reddish brown; Yield (65%, 0.375g); m. p.
360oC; IR (KBr/cm-1): 1604 νC=O, 3422 νOH, 1254 δOH, 1528 νC=C, 1111 νS=O, 512 νRu-O, 1384
1
νNO3; H NMR (400 MHz, DMSO-d6, 25oC, TMS): δ 6.90-8.70 (Ar-H), δ 3.60 (s, 6H, CH3,
DMSO-S), δ 2.50 (CH3, 6H, DMSO-O), δ 1.27 (s, OCH3); For RuNa2C36H36N2S2O18, ESI/MS
(979). Calcd.: C 44.13, H 3.68, N 2.86, S 6.54, Found C 44.38, H 3.82, N 2.86, S 6.38.
[Ru(DMSO)2(L2)2] 2NaNO3.H2O (M2R): Colour: Dark brown; Yield (55%, 0.278 g); m. p.
360oC; IR (KBr/cm-1): 1635 νC=O, 3435 νOH, 1174 δOH, 1508 νC=C, 1174 νS=O, 509 νRu-S, 1384
1
νNO3; H NMR (400 MHz, DMSO-d6, 25oC, TMS): δ 10.00 (s, OH), δ 7.00-8.32 (Ar-H), δ 3.00
(s, 6H, CH3, DMSO-S), δ 2.50 (s, 6H, DMSO, CH3) ; For RuNa2C34H32N4S2O19.; ESI/MS
(1009). Calcd: C 40.44, H 3.17, N 5.55, S 6.34, Found: C 40.37, H 3.23, N 5.38, S 6.57.
[Ru(DMSO)2(HL3)2](NO3)2.2H2O (M3R): Colour: Brownish black; Yield (70%, 0.365g); m. p.
360oC; IR (KBr/cm-1): 1605 νC=O, 3431 νOH, 1259 δOH, 1508 νC=C, 1111 νS=O, 509 νRu-S, 1384
6

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1
o
νNO3; H NMR (400 MHz, DMSO-d6, 25 C, TMS): δ 11.92 (s, OH), δ 13.14 (OH), δ 6.64-8.52
(Ar-H), δ 3.02 (d, DMSO-S), δ 2.82 (DMSO-O), δ 2.50 (N(CH3)2, 6H, d), δ 1.21 (s, OCH3); For
RuC38H46N4S2O16.; ESI/MS (979). Calcd.: C 46.58, H 4.70, N 5.72, S 3.27, Found C 46.28, H
4.46, N 5.54, S 3.42.
[Ru(DMSO)2(L4)2]2NaNO3.5H2O (M4R): Colour: Brown; Yield (65%, 0.378 g); m. p. 360oC;
1
IR (KBr/cm-1): 1686 νC=O, 3418 νOH, 1174 δOH, 1543 νC=C, 1091 νS=O, 548 νRu-S, 1384 νNO3; H
NMR (400 MHz, DMSO-d6, 25 oC, TMS): δ 7.17 -7.94 (Ar-H), δ 3.30 (s, 6H, DMSO-S), δ 2.5
(s, 6H, DMSO-O); For Ru NaC34H40N2 S2O19Cl.; ESI/MS (1060) Calcd.: C 38.53, H 3.78, N
2.64, S 3.02, Found C 38.25, H 3.74, N 2.34, S 3.47.
2.7 Computational details
In order to ascertain the nature of molecules and their electronic transition, density functional
theory (DFT) and time-dependent DFT calculations were performed. Optimized molecular
geometries were calculated using the B3LYP exchange-correlation functional [41, 42]. The
LANL2DZ basis set for Ru and 6-31G** basis set for other atoms were used for geometry
optimization. The optimized structures of the compounds were used for molecular orbital
analyses as well as for the time-dependent density functional theory (TD-DFT) calculations
using polarised continuum model (PCM) [40]. All the calculations were performed using
Gaussian 09 program [43].
R=-OCH3 (M1R);-NO2(M2R); -NMe2(M3R), -Cl(M4R)
Scheme 1. Synthetic routes for the ligands and metal complexes.
7

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3. Results and discussion
3.1 Optimized geometry of complexes
The optimized geometry of the metal complexes of Ru (II) are shown in Figure 1. The immediate
geometry around the Ru(II) in all the complexes are satisfied by two flavone ligands through
bidentate mode as well as by two DMSO ligands in monodentate mode there by leading to a
distorted octahedral geometry (Fig. 1). The calculated bond lengths and bond angles as presented
in Table 1 are comparable with the reported metrical parameters of the similarly structured
complexes [44]. In all the four complexes the Ru-O bond lengths are in good agreement with the
previously reported values [45].
Table 1. Optimized geometrical parameters for the metal complexes (bond length (Å) and bond
angle (°).
Parameters
Ru-S
M1R
M2R
M3R
M4R
2.251(2.282)
2.115 (2.134)
2.118 (2.087)
2.196
2.282 (2.282)
2.110 (2.134)
2.117 (2.087)
2.188
2.280(2.282)
2.115 (2.134)
2.197 (2.087)
2.127
2.255(2.282)
2.114 (2.134)
2.118 (2.087)
2.193
Ru-O1
Ru-O2
Ru-O3
2.120 (2.087)
2.132 (2.134)
120.32
2.116 (2.087)
2.127 (2.134)
118.30
2.119 (2.087)
2.133 (2.134)
120.00
2.120 (2.087)
2.130(2.134)
119.56
Ru-O4
Ru-O5
Ru-S-O6
Ru-O1-C
Ru-O2-C
Ru-O3-S
Ru-O4-C
Ru-O5-C
112.65
113.17
112.98
112.88
111.98
112.47
111.85
112.20
113.98
115.37
113.55
114.57
112.53
143.95
112.33
112.76
111.82
112.26
111.65
112.05
8

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M1R
M2R
M3R
M4R
Fig. 1. Perspective view of the optimized geometry of metal complexes (Colour legends: Red
Oxygen; Yellow Sulfur; grey carbon; dark green ruthenium and light green chlorine).
Table 2. IR (in cm-1) spectroscopic data for the ligands and Ru(II) complexes.
Compounds
HL1
νC=O
1684
1693
1692
1699
1604
1635
1605
1686
νO-H
3500
3366
3475
3423
3422
3435
3431
3418
δO-H
1283
1235
1273
1209
1254
1174
1259
1174
νC=C
1591
1520
1568
1566
1528
1508
1508
1543
νS=O
νRu-S
-
-
-
-
HL2
-
-
HL3
-
-
HL4
1111
1174
1111
1091
512
509
509
548
M1R
M2R
M3R
M4R
3.2 Infrared spectroscopy
The IR spectra of the ligands displayed ν(C=O) bands at 1684, 1693, 1692 and 1699 cm-1 for
HL1, HL2, HL3 and HL4, respectively, which on coordination to the Ru(II) center gets slightly
9

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shifted to lower energy to 1604, 1635, 1605 and 1686 cm-1 for M1R, M2R, M3R and M4R,
respectively there by confirming the coordination of C=O part of the ligand to ruthenium center.
In the IR spectra of M3R the major shift to lower energy (50 cm-1) observed for hydroxyl group
δOH indicated its coordination with the Ru(II) center. However, observed vibrations for the
hydroxyl group in other metal complexes (M1R, M2R and M4R) were due to lattice water [46]
1
which was further supported by mass spectrometry and H NMR spectroscopy. Furthermore, in
M1R the presence of peaks at 557 and 512 cm-1 corresponding to ν (Ru-O), supported that
hydroxyl group of flavone is coordinated. The coordinated DMSO molecule show band in the
range 1174-1091 cm-1 which was assigned to νS-O stretching of S-coordinated DMSO molecule
which is significantly shifted from the corresponding νS-O of free DMSO observed at 1050 cm-1
[46b]. Additionally presence of single band also indicates the presence of coordinated DMSO
molecule in trans-position. The appearance of peak at 1024, 1015, 1016 and 1015 cm-1 for M1R,
M2R, M3R and M4R, respectively is due to O-bonded DMSO molecule and lowering in
stretching frequency might be due to the lowering in bond order of S=O group. This is further
supported by Ru-S mode, which appears as two bands at 1111 and 1024 cm-1 [47]. The
magnitude of separation confirms a symmetric bidentate coordination for all Ru complexes. In
the solid state the nitrate stretch and deformation bands were also found in all complexes at
~1384 and 830 cm-1. The comparative IR spectroscopic data for the ligands and complexes are
presented in table 2.
3.3 NMR spectroscopy
1
The H NMR spectra display aromatic protons in the range 6.0 to 8.0 ppm and which indicates
the coordination of two ligands. The two sharp peaks in the range 3.39-2.50 ppm, confirmed
coordination of DMSO ligand through DMSO-S and DMSO-O, respectively [48]. In M1R, M2R
and M4R, the absence of signal at ~11.00 ppm indicates the deprotonations of OH proton in
corresponding flavone ligands. However, in M3R the appearance of two singlets at 11.92 and
13.14 ppm corresponding to the OH proton of flavones indicate that two flavones ligands are
1
coordinated to ruthenium center. The comparative H NMR spectroscopic data for the ligands
and complexes are presented in table 3.
10

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Table 3. 1H NMR (in ppm) spectroscopic data for the ligands and Ru(II) complexes.
Compounds
HL1
OH
10.67
10.86
13.35
10.57
-
Ar-H
DMSO-S
DMSO-O
6.99-7.73
7.16-7.50
7.14-7.50
7.22-7.73
6.90-8.70
7.00-8.32
6.64-8.52
7.17 -7.94
-
-
-
-
HL2
-
-
HL3
-
-
HL4
3.60
3.00
3.02
3.30
2.50
2.50
2.82
2.5
M1R
M2R
M3R
M4R
13.14
-
-
3.4 Mass spectrometry (ESI-MS)
In ESI-MS for M1R, of [Ru(DMSO)2(L1)2].2NaNO3.2H2O showed molecular ion peak at (m/z
980), ascribed at 979, further the fragmentation pattern showed loss of lattice water molecules
from coordination sphere, which can be ascribed to 943 for [Ru(DMSO)2(L1)2].2NaNO3 (m/z at
944), and loss of 2NaNO3 also ascribed to 775 (calc.) for [Ru(DMSO)2(L1)2] at (m/z 776). All
other complexes showed similar loss of water and sodium nitrate from outside of the
coordination sphere (Fig. S4). In the mass spectrum of M2R the peak corresponding to
[Ru(DMSO)2(L2)2].2NaNO3.H2O
was
observed
at
m/z
1010
(calcd.
1009);
[Ru(DMSO)2(HL2)2].2NaNO3 showed peak at m/z 990 (calcd. 991); the signal for
[Ru(DMSO)2(L2)2] observed at m/z 823 (calcd. 823); [Ru(L2)2] at m/z 667 (calcd. 667). The
mass spectrum for M3R [Ru (DMSO)2(HL3)2](NO3)2.2H2O showed peak at m/z 978 (calcd.
979);
[Ru(DMSO)(HL3)2] fragment was observed at m/z 663 (calcd. 662). In M4R
[Ru(DMSO)2(HL4)2].2NaNO3.5H2O shows peak at m/z 1059 (calcd. 1060);
[Ru(DMSO)2(HL3)2](NO3)2 fragment was observed at m/z 820 (calcd. 818);
a
[Ru(DMSO)2(L4)2].2NaNO3 fragment was observed at m/z 968 (calcd. 970); [Ru(DMSO)2(L4)2]
fragment was observed at m/z 801 (calcd. 802).
3.5 Electronic Absorption Spectroscopy
The electronic absorption spectra recorded in acetonitrile for all the complexes consisted of well-
defined bands in the range at 200-600 nm (Table 4). However, a number of significant
11

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differences between the ligand and metal complexes are arising due to the functionalization at 4′-
position of 3-hydroxy flavones (Fig. S3). The absorbance at 250-300 nm, can be assigned to π-π*
transition of the ligands. In addition, a series of strong ligand centered absorption were observed
which can be attributed to the DMSO ligands. Also the absorption observed in the range 428-550
nm is due to metal to ligand charge transfer. To gain further insight into the nature of electronic
absorption spectra TD-DFT calculations were carried out. The electron density distributions for
the selected frontier molecular energy levels for the complexes are shown in Fig. S5. In order to
locate the electronic transitions, HOMO must be localized on the donor site and LUMO should
be on the acceptor site. The electron densities of the highest occupied molecular orbitals are
mainly located with contribution of 50 and 52% respectively and the ligand character is about
35%. In the case of M2R, HOMO, HOMO-1 and HOMO-2 are entirely localized on the
Ruthenium centre and contribution of about 80% and 20% of the flavonol ligands. In the
LUMOs of M1R-M4R the contribution from the flavonols ligand was observed to be maximum.
Table 4. Electronic absorption spectroscopic data for the ligands and Ru(II) complexes.
Compound
Wavelength λmax (nm) ε0
203 (16400), 220 (2500), 237 (26400), 255 (30700), 270 (55300), 314
(53200), 355 (82500), 432 (24300).
HL1
222 (4000), 236 (4100), 256 (5200), 273 (19700), 290 (2500), 480(9600)
245 (2000), 277 (30800), 340 (34300), 455 (27800).
270 (7300), 310 (1800), 340 (1500), 368 (16900), 308 (27700), 347 (32000),
434 (65000).
HL2
HL3
HL4
220 (13300), 252 (3280), 269 sh (3360), 311 (16700), 330 (2900), 420
(2550), 430 (1600), 440 sh (1470).
M1R
M2R
310 (3200), 430 (2700), 510 sh (1200)
270 sh (2700), 400 sh (6900),
M3R
M4R
272 (32300), 310 (19500), 350(1800), 430 (4400), 463 (2200) shoulder.
3.6 Cell cytotoxicity
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The complex M1R was found to be most antiproliferative with IC50 value of 16 µM followed by
the activity of HL1, M2R, HL2, M3R and HL3 with the IC50 value of 17.2, 28.0, 29.5, 32.1 and
35.4 µM, respectively. However, M4R and HL4 were found to be least effective with the IC50
value of 36.2 and 38.4 µM, respectively (Table 5).
Table 5. Concentration for 50% inhibition, required to inhibit cancer cell proliferation by 50 %
of Ru(II) DMSO complexes along with p-substituted 3-hydroxy flavones.
HL1
M1R
HL2
M2R
HL3 M3R
HL4
M4R
Compounds
17.2
inhibition
16.0
29.5
28.0 35.4 32.1
38.4
36.2
Cell
IC50 (µM)
Similarly, dose dependent percentage antiproliferative activities for all compounds were
presented in the form of Mean±SE. The data were presented in three different ways of
comparisons such as between compounds and within concentration, within compounds and
between concentrations and among all the compounds (Table 6) and metal complexes compared
with standard drug cis-platin and other ruthenium complexes. HL1 and M1R were found to be
more effective (p<0.0001) as compared to all other compounds at each concentration while M1R
was most effective among all ligands and complexes. After the coordination of ligands with
Ru(II) metal, all the metal complexes were screened to be more active than their corresponding
ligands. The compounds HL1 and M1R exhibited maximum antiproliferative activities as
compared to the other all compounds in a dose dependent manner. The ligands showed activities
in the order HL1>HL2>HL3>HL4 and their Ru(II) complexes which were observed to be more
active than ligands follows the activity order M1R>M2R>M3R>M4R. Table 6 and Figure 2 are
representing the all possible comparisons in between compounds and their concentrations. The
compounds which revealed less than 50% antiproliferative activities can be considered suitable
DOSE in an in vitro analysis as comparable to IC50. The 5 and 10 μM dose of all the compounds
showed less than 50% antiproliferative activities. While at 20 μM HL2, HL3, HL4 and their
complexes exhibited less than 50% activities as well at 30 μM concentrations only HL3 and
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HL4 show less than 50% activities. All other doses of compound revealed cytotoxicity when
compared to IC50.
Table 6. Antiproliferative activities of all compounds and their comparative analysis
Compounds
Concentrations in µM(Mean±SE)
10 20 30
5
40
16.41±0.391 28.55±0.371,2 59.10±0.751 63.44±0.571,2 66.62±0.551,2
HL1
,2
,2
19.25±0.651 31.37±0.481 62.92±0.93
0.01* 0.004* 0.02*
64.66±0.52 67.91±0.641,2
0.16 0.18
M1R
p-value$
15.51±0.891 27.12±0.471 42.03±2.561 50.32±0.561,2 55.85±0.971,2
HL2
,2
18.33±1.251 30.87±0.821 43.55±0.841 52.16±0.791,2 61.06±0.631,2
M2R
0.12
0.007*
0.59
0.11
0.004*
p-value$
12.64±0.481 22.76±0.941 34.30±2.291 44.94±0.681,2 54.67±0.751,2
HL3
,2
16.11±0.821 26.83±0.711 39.22±1.401 47.72±0.481,2 58.91±0.591,2
M3R
,2
0.01*
0.01*
0.12
0.02*
0.004*
p-value$
HL4
8.54±0.431,2 16.75±0.971,2 27.20±0.441 38.92±1.251,2 52.53±0.691,2
,2
11.00±0.811 18.83±0.291 29.39±0.881 41.72±1.201,2 57.44±1.121
M4R
,2
0.04*
0.09
0.07
0.16
0.01*
p-value$
Unpaired t-test between compounds and within concentration, *Significant (p<0.05),
1Significant within compounds and between concentrations (p<0.0001), 2Significant (p<0.0001)
between HL1, HL2, HL3, HL4 and M1R, M2R, M3R, M4R
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The antiproliferative activity of HL1 was significantly higher (p<0.05) at concentration of 5 µM
(16.41±0.39), 10 µM (28.55±0.37) and 20 µM (59.10±0.75) than M1R as well as rest of the
ligands and complexes at same concentration. The activity of HL1 was significantly effective
(p<0.0001) in comparison to all other compounds. However, the activities corresponding to
M1R recorded at 5, 10, 20 µM concentrations were significantly higher as compared to HL1 and
other compounds (p<0.0001).
The effect of HL2 was significantly higher (p<0.05) at concentration 10 µM (27.12±0.47) and 40
µM (55.85±0.97) than M2R. The effect of HL2 and M2R was recorded significantly different
between all the concentrations (p<0.0001) (Table 6 and Figure 2). Similar observation was found
for HL4 and M4R (Table 5 and Figure 2). There was no significant difference between HL2,
HL3 and HL4 at concentration 5. However, HL1 (16.41±0.39) was significantly (p<0.001)
higher than HL4 (8.54±0.43) at concentration 5. Similar observation was found at concentration
10. No difference observed among M1R, M2R, M3R and M4R at concentration 5. The HL1
(59.10±0.75) values was significantly (p<0.001) higher than HL2 (42.03±2.56), HL3
(34.30±2.29) and HL4 (27.20±0.44) at concentration 20. Almost similar observation was found
at the concentration 30 and 40 for all the parameters and M1R, M2R, M3R and M4R. The
antiproliferative activities of the synthesized complexes show relatively lower IC50 values than
NAMI-A and other related ruthenium-DMSO complexes [49-51]. The results indicate that
ruthenium-DMSO flavone complexes may prove to be prospective candidates as anticancer
agents against breast cancer.
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Fig. 2. In vitro activity of ligands and their respective Ru (II) complexes against Human Breast
Cancer Cell Line MCF-7 measured by a 24 hr. MTT assay as described in the experimental
section.Values are expressed as means ± SE of at least three independent value.
3.7 Cell cycle assay
Analysis of cell-cycle phase distribution was carried out to study the antiproliferative mechanism
of most active compound HL1 and M1R on a single dose of 16µM. Figure3B shows the G1
checkpoint was interfered with both HL1 and M1R. However, G1 phase was dramatically
increased and S phase was reduced, when treated with M1R. These results indicated that M1R
induced G1 arrest by inhibiting DNA synthesis in S phase and cell apoptosis. G1 and G2 phases
of cell cycle are the major check points, have an important role in cell cycle progression [52-53].
Also, all activities were compared with previously reported antiproliferative activity of ofloxacin
and Ru (II) complexes on HeLa cell line [54-58] which indicated that Ru(II) complex containing
more aromatic ring will be exhibiting more effective antiproliferative activity [54]. In the same
vein, our results also indicated that the complexes reported herein show more effective
antiproliferative activity due to the presence of para-substituted aromatic rings on flavonols and
can be more persuasive drug than the previously reported drugs [59-60 ]. The antiproliferative
activity of M1R was investigated in MCF-7 breast cancer cells and was found to be more active
than cis-platin against GI50 of T-47D breast cancer [61-62].
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Fig. 3. Cell cycle distribution of MCF-7 cells treated with IC50 value of compounds L1 and M1R
for 24 h (A) Original cell cycle diagram (B) Quantitative distribution of percentage of cells in
different phases of cell cycle.
4. Conclusion
Investigation indicated that among the ruthenium(II) complexes of para-substituted-3-hydroxy
flavones reported herein, the complex M1R comprising of –OCH3 group was found to be most
antiproliferative with IC50 value of 16 µM followed by the activity of HL1, M2R, HL2, M3R
and HL3. M4R and HL4 were found to be least effective. Analysis of cell-cycle phase
distribution was carried out to study the anti-proliferative mechanism of most active compound
HL1 and M1R on a single dose of 16 µM. They show the G1 check point was interfered with
both HL1 and M1R. This inhibits G1 arrest by inhibiting DNA synthesis in S Phase and cell
apoptosis. It can be concluded that the ruthenium(II) complexes having para- substituted 3-
hydroxy flavones with DMSO as ancillary ligands can become potential candidate as anticancer
agents in future. The antiproliferative activities of the synthesized complexes show relatively
lower IC50 values than NAMI-A and other related ruthenium-DMSO complexes. The results
indicate that ruthenium-DMSO flavone complexes may prove to be prospective candidates as
anticancer agents against breast cancer.
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5. Acknowledgements
Ashok. K. Singh would like to thank to UGC major research project No.F-39-701/2010(SR) for
financial assistance. Authors are also grateful to the Department of Chemistry, Lucknow
University, Lucknow and Director, CDRI, Lucknow, India and for support to carry out the study.
Author G.S. would like to thank UGC for financial support.
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References:
[1]
B. Lipper, E.; VHCA: Zurich, Cisplatin: Chemistry and Biochemistry of Leading
Anticancer Drugs. 1999.
[2]
[3]
E. Wong, C.M. Giandomenico. Chem. Rev. 99 (1999) 2451-2466.
(a) P.J. Loehrer,; L.H. Ann. Einkorn.Intern. Med., 100 (1984) 731; (b) M. Sun, Science.,
222 (1983) 145.
[4]
[5]
[6]
(a) F.T. Garzon, M. R. Berger, B. K. Kipper. D. Schmahl, Pharmacol,19 (1987) 347; (b)
M. J. Clarke. Metal ions in Biological System, 11 (1980) 231-283.
(a) M. J. Clarke, F. Zhu, D.R. Frasca. Chem. Rev. 99 (1999) 2511 ; (b) M. J. Clarke.
Coord. Chem. Rev., 236 (2003) 209 .
(a) G. Mestroni, E. Alessio, G. Sava, S. Pacor, M. Cilicia, B. K. Kipper.Ed., VCH:
Wenham (Germany), 1993, 157; (b) G. Medtronic, E. Alessio, G. Sava, S. Pacor, M.
Coluccia, A. Boccarelli. Metal-Based Drugs, 1 (1994) 41; (c) H. Huang, P. Zhang, Y.
Chen, K. Qiu, C. Jin, L. Ji, DOI: 10.1039/c6dt01270a; (d) E.M. Njogu, B. Omondi, V. O.
Nyamori, H. Chao, J. Coord. Chem., 68 (2015) 3389–3431; (e) R. Prajapati, V.K.
Yadav, S.K. Dubey, B. Durham, L. Mishra, Ind. J. Chem, 47 (2008) 1780-1786.
P. S. K Wong, Y. Chun-Wing, C. F. Y. Steve, Y. H. Ping. Biochem. Biophys. Res.
Comm., 363 (2007) 235-240.
[7]
[8]
[9]
A. Bergamo, G.Sava. Dalton Trans. 40 (2011) 7817 - 7823.
R. Prajapati, S.K. Dubey, R. Gaur, R. K. Koiri, B.K. Maurya, S. K. Trigun, L. Mishra
Polyhedron, 29 (2010) 1055-1061.
[10] I. P. Evans, A. Spencer, G.Wilkinson. J. C. S. Dalton Trans., (1973) 204-209.
[11] L. T. Lemiesz . Acta. Biochim. Pol., 52 (2004) 199.
[12] A Bergamo, G. Stocco, C. Casarsa, M.Cocchettio, E. Alessio, B. Serli, S. Zorzet, G.
Sava. Int. J. Oncol , 24 (2004) 373.
[13] G. Sava, R.Gangliardi, A. Bergamo, E. Alessio, G. Mestroni. Anti Cancer Res., 19
(1999) 969-972.
[14] B. K. Kipper, U. M. Henn, M. R. Juhl,R. Berger, F.E.Niebi, A. Wagner.Prog.
Clin.Biochem. Med., 10 (1989) 41-69.
[15] (a) J. Siemiatycki, L. Richardson, K. Straif, B. Latreille, R. Lakhani, S. Campbell, M.
C. Rousseau, P. Boffetta. Environ. Health Perspect., 112 (2004) 1447-59; (b) C. G.
19

ACCEPTED MANUSCRIPT
Hartinger, S. Zorbas-Seifried, M. A. Jakupec, B. Kynast, H. Zorbas, B. K. Kipper, J.
Inorg. Biochem., 100 (2006) 891-904.
[16] A. D. Lima, F. D. Castropereira, C. Costa, A. D. Santanabragabarbosaribeiro,
Luizalfredopavanin, W. D. S. D. Paulasilveira-Lacerda. J. Biosci ., 35 (2010) 371–378.
[17] M. J. Menezes, S. Manjrekar, V. Pai, R. E.Patre. Ind. J. Chem., 48A (2009) 1311-1314.
[18] O.Novakova, J. Kasparova, O. Vrana, P. M.VanVliet, J. Reedijk, V.Brabee.
Biochemistry, 34 (1995) 12369-12378.
[19] B. K.Keppler, D. Wehe, H. Endres, W. Rupp. Inorg. Chem., 26 (1987) 844-896.
[20] B. K. Kepper, W. Rupp, U. M. Juhl, H. Endres, R. Nieu, W.S. Balzer.Inorg. Chem., 26
(1987) 4366 – 4370.
[21] Z. Nowakowska. Eur. J. Med. Chem., 42, (2007) 125-137.
[22] B. P Bandgar, S. S. Gawande, R. G. Bodade, J. V. Totre, C.N. Khobragade.Bioorg. Med.
Chem., 18 (2010) 1364-1370.
[23] N. Gais,M.M. Rahman, M.A.Rashid, H. Kashino, K. Nageswara, T. Nakata. Fitoterapia,
67 (1996) 554.
[24] M. Hecker, C. Presis, P. Klemm, B. R. Busser. J. Pharmacol , 118 (1996) 2178.
[25] R. J. Fishkin, J. T.Winslow. Psychopharmacology (Berl.), 132 (1997) 335.
[26] F.L. Pearce, A.D. Befs, J. Blenenstock. J. J. Allergy Clin. Immunolgy, 73 (1984) 819.
[27] S. Bhatnagar, S. Sahi, P. Kackar, S. Kaushik, M.K. Dave, A. Shukla, A. Goel. Bioorg.
Med. Chem. Lett., 20 (2010) 4945-4950.
[28] H.Goker,S. Ozden, S. Yildiz, D. W. Boykin.Eur. J. Med. Chem. 40, (2005) 1062-1069.
[29] G. B. Liu, J.L. Xu, M. Geng, R. Xu, R. R. Hui, J.W. Zhao, Q. Xu, H.X. Xu, J.X. Li.
Bioorg. Med. Chem., 18 (2010) 2864-2871.
[30] S. Habtemariam.J. Nat. Products., 60 (1997) 775.
[31] Y. C.Kao, C.Zhou, M. Sherman, C. A. Loughton, S. Chen. Environ. Health Perspect.,
106 (1998) 85.
[32] Y. Wang,C.T. Ho, T. Yoshikawa.Forum Nutr. Basel, Karger. 61 (2009) 64–74.
[33]
[34]
J. Velisek, J. Davidek, K. Cejpek, Czech. J. Food Sci., 26 (2008) 73–98.
Hirao, Synthesis, 12 (1982) 1076.
[35] Vogel’s Text book of practical organic chemistry, 5th Edition, Solvents and reagents (The
purification of common organic solvents, (1987) 264-319.
20

ACCEPTED MANUSCRIPT
[36] A. Mohammad, S. Singh, A. Sharma, N. Verma, Indus. Engin. Chem. Res., 52 (2013)
4672-4682.
[37] P.Zhang, J. Chen, Y. Liang, Acta Biochim Biophys Sin (Shanghai), 42 (2010) 440-449.
[38] Y. Hoshino, T. Oohinata. Bull. Chem. Soc. Jap., 59 (1986) 2351.
[39] T.S. Wheeler. Org. Syn. Coll., 44 (1963) 78.
[40] B. Chakravarti, R.Maurya, J.A. Siddiqui, H.K. Bid, S. M.Rajendran, P.P.Yadav,
R.J.Konwar.Ethnopharmacol, 142 (2012) 72-79.
[41] A. D. Becke. J. Chem. Phys. 98 (1993) 5648.
[42] V. Barone, M. Cossi and J. Tomasi. J. Comput. Chem. 19 (1998) 404-417.
[43] M. J. Frisch, G. W. Trucks, H. B. Schlegel, G. E. Scuseria, M. A. Robb, J. R. Cheeseman,
J. A. Montgomery, T. Vreven Jr., K. N. Kudin, J. C. Burant, J. M. Millam, S. S. Iyengar,
J. Tomasi, V. Barone, B. Mennucci, M. Cossi, G. Scalmani, N. Rega, G. A. Petersson, H.
Nakatsuji, M. Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T.
Nakajima, Y. Honda, O. Kitao, H. Nakai, M.; Klene, X. Li, J. E. Knox, H. P. Hratchian,
J. B. Cross, V. Bakken, C. Adamo, J. Jaramillo, R. Gomperts, R. E. Stratmann, O.
Yazyev, A. J. Austin, R. Cammi, C. Pomelli, J. W. Ochterski, P. Y. Ayala, K.
Morokuma, G. A. Voth, P. Salvador, J. J. Dannenberg, V. G. Zakrzewski, S. Dapprich,
A. D. Daniels, M. C. Strain, O. Farkas, D. K. Malick, A. D. Rabuck, K. Raghavachari, J.
B. Foresman, J. V. Ortiz, Q. Cui, A. G. Baboul, S. Clifford, J. Cioslowski, B. B.
Stefanov, G. Liu, A. Liashenko, P. Piskorz, I. Komaromi, R. L. Martin, D. J. Fox, T.
Keith, M. A. Al-Laham, C. Y. Peng, A. Nanayakkara, M. Challacombe, P. M. W. Gill, B.
Johnson, W. Chen, W. M. Wong, C. Gonzalez, J. A. Pople, Gaussian, Inc., Wallingford
CT, 2004.
[44] I. Bratsos, B. Erli, E. Zangrando, N. Katsaros, E. Alessio. Inorg. Chem., 46 (2007)
975-992.
[45] Y. Hoshino, T. Oohinata. Bull. Chem. Soc. Jap., 59 (1986) 2351.
[46] K. Nakamoto, Infrared and Raman spectra of inorganic compounds.4th Edition, 227-228
(1986)
[47] K. Nakamoto, Infrared and Raman spectra of Inorganic compounds.4th Edition, 270
(1986).
21

ACCEPTED MANUSCRIPT
[48] I. Bratsos, B. Erli, E. Zangrando, N.Katsaros, E. Alessio. Inorg. Chem., 46 (2007) 975-
992.
[49] K. K. Jovanovick, N. Gligorijevic, R. Gaur, L. Mishra, S. Radulonic. J. Boun., 21 (2016)
482-4890.
[50] S. Pillozzi, L. Gasparoli, M. Stefanini, M. Ristori, M. D Amico, E. Alessio, F. Scaletti, A.
Becchetti, A. Arcangeli, B. Messori, Dalton Trans., 43 (2014) 12150-12155.
[51] R. Prajapati, S.K. Dubey, R. Gaur, R. K. Koiri, B.K.Maurya, S.K. Trigun, L. Mishra,
Polyhedron, 29 (2010)1055-1061.
[52] I. B. Chakravarti, R.Maurya, J.A. Siddiqui, H.K. Bid, S. M.Rajendran, P. P. Yadav, R. J.
Konwar. Ethnopharmacol, 142 (2012) 72-79.
[53] M.N. Patel, D.S. Gandhi, P.A. Parmar, H.N. Joshi, J. Coord. Chem, 65 (2012) 1926-
1936.
[54] C. Wang, Q. Wu, Y. Zeng, D. Huang, C. Yu, X. Wang, W. Mei, J. Coord. Chem., 68
(2015) 1489-14999.
[55] E. Njogu, B. Omondi, V. Nyamori., J. Coord. Chem., 68 (2015) 3389-3431.
[56] X. Feng, W. Chen, B.Xiang, J. Coord. Chem., 69 (2016) 1551-1558.
[57] M. Shilpa, C. Shobha Devi, P. Nagababu, J.N.L. Latha, R. Pallela, V. R. Janapala, K.
Aravind, S. Satyanarayana, J. Coord. Chem., 66 (2013) 1661.
[58]
J. Sun, W.X. Chen, X.D. Song, X.H. Zhao, A.Q. Ma, J.X. Chen, J. Coord. Chem., 68
(2015) 308.
[59] S. H. Liu, J.Wei, H. H. Xu, Y. Wang, Y. M. Liu, J. B. Liang, G. Q. Zhang, D.H. Cao, Y.
Y. Lin, Y. Wu, Q.F. Guo, spectrochim. Acta part A 161 (2016) 77-82
[60] M. Voicescu, S. Lonescu, C. L. Nistor, spectrochim. Acta part A 170 (2017) 1-8
[61] Z. Mendoza, P. Lorenzo, M. Serrano-ruiz, E. Martin-Batista, J.M.Padron, F. Scalambra,A.
Romerosa, Inorg. Chem., DOI: 10.1021/acs.inorgchem.6b01207
[62] A. K. Singh, G. Saxena, S. Dixit, Hamidullah, Sachin. K. Singh, S. K. Singh, M. Arsad,
R. Konwer, J. Mol. Strc. 111(2016) 90
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Synthesis, Characterizations and Biological evaluation against breast cancer of Ruthenium
Flavanol Complexes
Ashok Kumar Singha*, Gunjan Saxenaa, Sahabjadab and M. Arshadb
aDepartment of Chemistry, University of Lucknow, Lucknow – 226 007 INDIA
bDepartment of Zoology, University of Lucknow, Lucknow – 226 007 INDIA
*Corresponding author: Ashok Kumar Singh; Phone: +91-9919663856;
E-mail address: singhaks3@rediffmail.com
Graphical abstract
The Ru(II)-DMSO complexes containing different 4-substituted 3-hydroxy flavones synthesized
which showed significant activity against breast cancer cell line, MCF-7.
M1R
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Research highlights:
 Ru(II)-DMSO flavonol complexes synthesized.
 The complexes were characterized using integrated physico-chemical technique
 The complexes were tested against breast cancer cell lines.
24