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Dedicated

To

My Loving Grand Mother (late) & Parents (late)

CONTENTS

ACKNOWLEDGEMENTS i-ii

ABSTRACT iii-iv

Tables List v

Figures List vi-ix

Abbreviations List x-xi

Chapter 1 INTRODUCTION 1-37

1.1 Cisplatin history 2

1.2 Cisplatin mode of action 3

1.3 Cisplatin toxicity 4

1.4 Resistance to cisplatin 6

1.4.1 Pre-binding resistance 7

1.4.1.1 Decrease cisplatin accumulation 7

1.4.1.2 Inactivation by thiol containing molecules 10

1.4.2 Post-binding resistance 10

1.5 Monofunctional platinum(II) complexes 11

1.5.1 Development of monofunctional platinum complexes 11

1.5.2 Highly potent monofunctional platinum complexes 15

1.6 Monofunctional platinum complexes with alternative cell kill mechanisms 16

1.6.1 Autophagy 17

1.6.2 Necrosis 19

1.6.3 Paraptosis 20

REFERENCES 24-37

Chapter 2 EXPERIMENTAL 38-74

2.1 Materials 38

2.2 Instrumentation 38

2.3 General procedure for synthesis of the ligands 39

2.4 General procedure for the synthesis of heteroleptic Pt(II) complexes 43

2.5 Computational studies 68

2.6 DNA-binding using UV-visible spectroscopy 69

2.7 DNA-binding using viscosity measurements 69

2.8 DNA-binding using cyclic voltammetry 70

2.9 Cytotoxic potential against HepG2 cancer cell line 70

2.10 Cytotoxic potential against five cancer cell lines 71

2.11 Anticancer activity against LU and MCF-7 cancer cell lines 71

2.12 Cleavage of plasmid DNA 72

REFERENCE 73-74

Chapter 3 RESULTS AND DISCUSSION 75-122

3.1 FT-IR 75

3.2 1H NMR 75

3.3 13

C NMR 76

3.4 31

P NMR 77

3.5 Anticancer study 90

3.5.1 Anticancer activity against HepG2 cell line 90

3.5.2 Anticancer activity against five different cancer cell lines 92

3.5.3 Anticancer activity against LU human lung carcinoma, MCF-7 human breast adenocarcinoma 94

3.6 DNA-Interaction study 95

3.6.1 UV-Visible spectroscopy 96

3.6.2 Viscometry 103

3.6.3 Cyclic voltammetry 106

3.7 Plasmid DNA cleavage 110

3.8 Chloride exchange 112

REFERENCES 119-122

Chapter 4 CRYSTALLOGRAPHIC ANALYSIS AND DFT STUDIES 123-144

4.1 Single crystal X-ray analysis 123

4.2 Crystal packing 128

4.3 Theoretical studies 134

REFERENCES 142-144

CONCLUSION 145-146

LIST OF PUBLICATIONS 147

iii

ABSTRACT

In the present study, 42 new heteroleptic Pt(II) dithiocarbamates were synthesized by the

reaction of PtCl2 with NaS2C-R (where R = 4-(4-methoxyphenyl)piperazine (L-1), 4-(2-

furoyl)piperazine (L-2), 4-diphenylmethylpiperazine (L-3), 4-(4-nitrophenyl)piperazine

(L-4), 4-(2-hydroxyethyl)piperazine (L-5), 4-benzylpiperazine (L-6), 4-(4-

hydroxyphenyl)piperazine (L-7), morpholine (L-8), piperidine (L-9) and 4,4ʹ

trimethylenedipiperidine (L-10) and organophosphines in chloroform-methanol mixed

solvents system. Various organophosphines used were tris-p-flourophenylphosphine, tris-

p-chlorophenylphosphine, diphenyl-p-tolylphosphine, tri-p-tolylphosphine, tris-p-

methoxyphenylphosphine and 1,4-bis(diphenylphosphino)butane. These complexes were

characterized by different analytical techniques namely elemental analysis, FT-IR,

multinuclear (1H,

13C and

31P) NMR and single crystal X-ray analysis along with DFT

calculations. Based upon results, monofunctional complexes showed pseudo square

planar geometry around platinum atom with two cis sites occupied by dithiocarbamate

moiety forming four-membered chelate ring (PtS2C) and the remaining two by chloride

and organophosphine. However, in bis-orgnophosphine complexes the latter cis positions

are occupied by the phosphorous atoms.

The complexes were examined for their in vitro cytotoxic potential against HepG2 human

hepatocellular carcinoma,

LU human lung carcinoma, MCF-7 human breast

adenocarcinoma, MDA-MB-231 human breast adenocarcinoma, Hepa-IcIc7 mouse liver

hepatoma and PC-3 human prostate adenocarcinoma by sulforhodamine B (SRB) cellular

protein-staining and MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

methods using doxorubicin and cisplatin as standard drugs. Generally, the

monofunctional complexes showed high activity against different cancer cell lines than

both the standard drugs.

Interaction of the representative Pt(II) complexes with DNA was examined by UV-Vis

spectroscopy, viscometry and cyclic voltammetry. The hyperchromic effect observed for

the studied complexes is an indication of the electrostatic interaction, a consequence of

distortion in the metal coordination core via the Pt-Cl bond dissociation. This fact was

iv

further supported by viscometric and cyclic voltammetric results. Furthermore, in order

to get concert evidence regarding the labile nature of Pt-Cl bond, substitution of chloride

by various ligands was followed by FT-IR, 1H and

31P NMR. Plasmid DNA cleavage

studies by agarose gel electrophoresis revealed that these complexes have the ability to

convert super coiled DNA to nicked circular DNA.

v

Tables List

Table Title Page No

1.1: Morphology changes of cells via apoptosis, autophagy, necrosis, and paraptosis cell death pathway 19

3.1: 1H NMR data of the ligands (L1 and L2) and their Pt(II) complexes (1-10) 80





3.6: 13

C NMR and 31

P NMR data of the ligands (L1 and L2) and their Pt(II) complexes (1-10) 85

3.7: 13

C NMR and 31


3.8: 13

C NMR and 31


3.9: 13

C NMR and 31


3.10: 13

C NMR and 31


3.11: IC50 values of the selected Pt(II) complexes against HepG2 cell line 91

3.12: IC50 values (µM) of the complexes (1, 2 and 6) against five different cancer cell lines 93

3.13: IC50 values (µM) of the complexes against Lu and MCF-7 cancer cell lines 95 95

3.14: Binding constants (K) and Gibb’s free energies (ΔG) of selected Pt(II) complexes based upon

UV- visible spectroscopic data 97

3.15: 31

P NMR peaks (ppm) of the representative complexes in the presence of different exchangeable

ligands 115

4.1: Crystal data and structure refinement for the complexes 1, 2 and 6 125

4.2: Crystal data and structure refinement for the complexes 25 and 29 126

4.3: Selected bond lengths of the complexes 1, 2, 6, 25 and 29 127

4.4: Selected bond angles of the complexes 1, 2, 6, 25 and 29 127

4.5: Various weak non-covalent interactions (Å) forming 1D and 3D networks for crystal packing 133

4.6: Comparison of the selected bond lengths by theoretical and X-ray calculations for the complexes 1, 2,

6 and 25 136

4.7: Comparison of the selected bond angles by theoretical and X-ray calculations for the complexes 1, 2, 6

and 25 136

4.8: Pt···H and Pt···C distances (Å) of the optimized structures of the representative complexes 137

4.9: NBO atomic charges of representative complexes from natural population analysis 140

4.10: Molecular properties of representative complexes 141

vi

Figures List

Figure Title Page No

1.1: Structures of globally used platinum anticancer drugs. 3

1.2: Different pathways of cisplatin before and after it enters the cell. 4

1.3: Pathophysiological events in cisplatin nephrotoxicity. 5

1.4: Affected parts of cochlea during ototoxicity. 6

1.5: Schematic diagram of hCtr1 structure, showing 190 amino acid residues, each represented by a circle.

The extracellularly located N-terminus, and intracellularly located C-terminus are indicated. The N-

terminal region contains two each His-rich (yellow) and Met-rich (red) motifs. The C-terminal His/Cys

residues are also indicated. 8

1.6: Schematic diagram showing transport of copper and cisplatin in and out of the cell. 9

1.7: General structure of monofunctional platinum(II) complexes. 11

1.8: Structure of ethidium-containing monofunctional complex. 12

1.9: General structure of cationic monofunctional complexes active in the S180a, P388 and L1210 screen. 13

1.10: Structures of monofunctional complexes 2-21. 14

1.11: Structures of bifunctional (22-24) and monofunctional (25-30) Pt(II) complexes with increasing

hydrophobicity and enhanced anticancer activity. 16

1.12: Monofunctional Pt(II) complexes that form Pt-DNA crosslink to induce apoptosis of the cancer cells. 17

1.13: The fate of cancer cells when they are induced by stress. 18

1.14: The possible cell death pathways for cancer cells bearing wild-type p53 or mutant p53 when treated

with cisplatin and monofunctional Pt(II) compound. 20

1.15: Monofunctional Pt(II) compounds suppress Akt and mTOR pathway to induce autophagy. 21

1.16: Monofunctional Pt compounds that did not interact with DNA and exhibit an alternative cell death

pathway. 21

1.17: General structure of the complex having two inert ligands and a labile ligand 22

3.1: Resonance forms of the dithiocarbamic-NCSS moiety. 75

3.2: The 1H NMR spectrum of the representative ligand (L1). 77

3.3: The 1H NMR spectrum of the representative monofunctional Pt(II) complex (2). 78

3.4: The 13

C NMR spectrum of the representative ligand (L1). 78

3.5: The 13

C NMR spectrum of the representative monofunctional Pt(II) complex (2). 79

3.6: The 31

P NMR spectrum of the representative monofunctional Pt(II) complex (2). 79

3.7: Comparison of IC50 (µM) values of the representative compounds against HepG2 cell line. 91

3.8: Comparison of IC50 (µM) values of the representative compounds against HepG2 cell line. 92

3.9: Comparison of IC50 (µM) values of 1, 2 and 6 against five different cancer cell lines. 93

3.10: Structure of 1 showing to have hydrogen bonding ability due to presence of flouro moieties. 94

3.11: Structure of 6 showing to have hydrogen bonding ability due to presence of flouro moieties. 94

vii

3.12: Absorbance of 35µM complex (1) in the absence (a) and presence of (b) 5µM, (c) 15µM, (d) 20µM,

(e) 25µM and (f) 30 µM DNA. The inset graph represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for

calculation of binding constant (K) and Gibb’s free energy (ΔG). 98

3.13: Absorbance of 25µM complex (2) in the absence (a) and presence of (b) 5µM, (c) 10µM, (d) 15µM,

(e) 20µM and (f) 25µM DNA. The inset graph represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for


3.14: Absorbance of 25µM complex (6) in the absence (a) and presence of (b) 3µM, (c) 6µM, (d) 9µM, (e)

12µM, (f) 15 µM and (f) 18µM DNA. The inset graph represents the plot of Ao/A-Ao vs. 1/[DNA]

(µM)-1

for calculation of binding constant (K) and Gibb’s free energy (ΔG). 100

3.15: Absorbance of 60 µM complex (16) in the absence (a) and presence of (b) 3µM, (c) 6µM, (d) 9µM,

(e) 12µM, (f) 15 µM, (g) 18µM and (h) 21µM DNA. The inset graph represents the plot of Ao/A-Ao

vs. 1/[DNA] (µM)-1


3.16: Absorbance of 100 µM complex (18) in the absence (a) and presence of (b) 5µM, (c) 10µM, (d)

15µM and (e) 20µM DNA. The inset graph represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for


3.17: Absorbance of 100 µM complex (19) in the absence (a) and presence of (b) 5µM, (c) 10µM, (d)

15µM, (e) 20µM, (f) 25µM and (g) 30µM DNA. The inset graph represents the plot of Ao/A-Ao vs.

1/[DNA] (µM)-1


3.18: Effects of increasing amounts of complexes 1-5 on the relative viscosities of CT-DNA at room

temperature, [DNA] = 100µM, r = [complex]/[DNA]. 104







3.22: Representative cyclic voltammogram of 1mM complex 2 in the absence of DNA (red) and in the

presence of 80 µM DNA (black) in DMSO with 0.5 M TBAP as supporting electrolyte at 50 mVs-1

scan rate. 106

3.23: Cyclic voltamograms of 1 mM complex 2 with 0.5 M TBAP as supporting electrolyte in the absence

(red) and presence of 20 μM DNA (green), 40 μM DNA (black), 60 μM DNA (blue) and 80 μM

DNA (orange) showing a decrease in current. 107

3.24: Representative plot of log (I/Io-I) versus log (1/[DNA]) for determination of binding constant of

complex (2). 107

3.25: Representative cyclic voltammogram of 1mM complex 2 at different (50-500 mVs-1

) scan rates in

DMSO with 0.5 M TBAP as supporting electrolyte. 108

viii

3.26: Representative cyclic voltammogram of 1mM complex 2 in the presence of 20 µM DNA at different

(50-500 mVs-1

) scan rates in DMSO with 0.5 M TBAP as supporting electrolyte. 108

3.27: Representative plot for determination of diffusion coefficients of free drug (2) and drug-DNA adduct. 109

3.28: Effect of complexes 4, 5, 7, 8, 9, 10, 11, 12, 13, 14, 18, 19 and 28 (50 µg/ml) on the cleavage of

pUC19 DNA(12.5 μg/ml) in Tris–HCl buffer (0.1 M, pH 7.4) at 37 °C after incubation for 6 h. Form

I = suppercoiled DNA, Form II = nicked circular DNA. 110

3.29: Effect of complexes 1, 2 and 6 (50 µg/ml) on the cleavage of pUC19 DNA (12.5 μg/ml) in Tris–HCl

buffer (0.1 M, pH 7.4) at 37 °C after incubation for 6 h. Form I = suppercoiled DNA, Form II =

nicked circular DNA. 110

3.30: Effect of concentration of complexes (a) 1 and (b) 4, on the cleavage of pUC19 DNA(12.5 μg/ml) in

Tris–HCl buffer (0.1 M, pH 7.4) at 37 °C after incubation for 6 h. 111

3.31: Comparison of IR spectra of representative complex 32 and its product 32a showing appearance of

new peaks due to thiocyanate attachment via N or S atom. 112

3.32: Comparison of 1HNMR spectra of representative complex 32 and its product 32a showing no change

in the peak positions. 112

3.33: Comparison of 31

P NMR spectra of the representative complex 32 and its product 32a showing small

upfield shift. 113

3.34: 31

P NMR spectra of the complex 32 (25 mM) in the presence of equal molar solution of iodide ions

recorded after 10, 20 and 30 min. of mixing. 114

3.35: 31

P NMR spectra of the complex 34 (25 mM) in the presence of excess of thiourea recorded after 2, 4

and 6 hours of mixing. 114

3.36: 31

PNMR spectra of the complex 32 (10 mM) in the presence of equal molar solution of

diethyldithiocarbamate recorded after different intervals of time. 115

3.37: Spectrum of change of absorbance with respect to time using diethyldithiocarbamate (110 µM) and

complex 32 (10 µM). Insight is the graphs between Ln ∆Abs verses time. 116


complex 32 (10 µM). Inset is the graphs between Ln ∆Abs verses time. 117


complex 32 (10 µM). Inett is the graphs between Ln ∆Abs verses time. 117


complex 32 (10 µM). Insight is the graphs between Ln ∆Abs verses time. 118

3.41: Graph between apparent rate constants vs different concentrations of DT (diethyldithiocarbamate). 118

4.1: Structures of monofunctional complexes (1, 2, 6 and 25). 124

4.2: Structure of complex (29) showing pseudo-square planner geometry. 124

4.3: Crystal structures of the complexes (1, 2, 6, and 25) showing steric hindrance from aromatic C-H

groups of organophosphine, C-Pt distance (black), H-Pt distance (red). 130

4.4: 1D-supramolecular chains in 3D-crystal packing of 1, 2, 6 and 25. 131

ix

4.5: The 3D-crystal packing of 1 along b-axis (a), 2 along b-axis (b), 6 along a-axis (c) and 25 along b-axis

(d). 132

4.6: The crystal packing of 29, 1D-packing through methanol bridging (a), 3D-packing along a-axis (b). 134

4.7: Comparison of single crystal XRD structure (a) and optimized structure (b) of the representative

complex 2. 135

4.8: Optimized structures of the complexes (1 and 2) showing steric hindrance from aromatic C-H groups

of organophosphine, C-Pt distance (black), H-Pt distance (red). 137

4.9: Structures of representative monofunctional complexes having atomic charge at each atom

representing from color of each atom, assessed from natural population analysis. Change from green

or red to black represents change of atomic charge (positive or negative) towards neutrality. 138

4.10: Frontier orbitals of the representative complexes showing energy difference between HOMO and

LUMO orbitals. 139

1

Chapter 1

INTRODUCTION

Cancer or malignant tumor/neoplasm is a group of diseases involving uncontrolled

growth of abnormal cells in one body part and propagate to other organs with the passage

of time [1]. Cancer is caused by changes to genes that control the way our cells function,

especially how they grow or divide. There are mainly four types of genes, namely proto-

oncogenes (involved in normal cell growth), tumor suppressor genes (also involved in

controlling cell growth), DNA repair genes (involved in fixing damaged DNA) and self-

destruction genes (genes that direct a cell to die), which are affected and cause cancer.

There are over 100 diverse known cancers affecting human’s life [1], thus responsible for

14.6% of the total human deaths according to the world health organization (WHO) [2].

Chemotherapy, surgery, radiotherapy, hormone therapy, immunotherapy, biological

therapy, photodynamic therapy, hyperthermia, bone marrow and stem cell transplant or

their combination therapies are the main remedies which are being used to control

malignancy. Among these cancer healing strategies, chemotherapy, use of anticancer

drugs to treat cancerous cells, is being used from many years. There are variety of such

chemotherapeutics each with its own mechanism of action such as alkylating agents,

antitumor antibiotics, antimetabolites, antimicrotubule agents and platinum complexes.

Alkylating agents alter structure of the guanine base of DNA via alkyl substitution,

exemplified by nitrogen mustard (mechlorethamine, cyclophosphamide, chlorambucil,

melphalan and ifosfamide), nitrosureas (carmustine, lomustine and streptozocin),

alkylsulfonates (busulfan) etc. However, carcinogenicity to normal cells [3], cell-cycle

non-specific nature and high activity in resting phase of the cell are associated adverse

effects of this class of drugs.

Antitumor antibiotics are natural products produced by species of soil fungus

Streptomyces. These drugs are considered cell-cycle specific and interrupt DNA and

ultimately cells replication. Such drugs include anthracyclines (doxorubicin,

daunorubicin, epirubicin), chromomycins (dactinomycin, plicamycin) and miscellaneous

(mitomycin, bleomycin). Antimetabolites, chemicals having structural resemblance to

http://en.wikipedia.org/wiki/Malignancy

http://en.wikipedia.org/wiki/Neoplasm

http://en.wikipedia.org/wiki/Neoplasm

http://en.wikipedia.org/wiki/Cell_growth

http://en.wikipedia.org/wiki/Alkylating_agent

http://en.wikipedia.org/wiki/Guanine

http://www.webmd.com/hw-popup/dna

2

metabolites that they interfere with [4], induce cell death during the S phase of the cell

growth either incorporated into RNA and DNA or inhibit enzymes needed for nucleic

acid production [5]. Antimetabolites include methotrexate, fluoropyrimidines (e.g. 5FU,

capecitabine), cytocine arabinose (e.g. cytarabine), gemcitabine etc. Antimicrotubule

agents {taxanes (docetaxel and paclitaxel), Vinca alkaloids (vincristine, vinorelbine, and

vinblastine), and estramustine phosphate} interfere with microtubules (cellular structures

that help move chromosomes during mitosis) and obstruct cell growth by arresting

mitosis [6-8]. Other way to treat cancer involves the use of monoclonal antibodies that

bind only to cancer cell-specific antigens and induce an immunological response against

the target cancer cell. Monoclonal antibodies include alemtuzumab, bevacizumab,

trastuzumab etc [9].

1.1 Cisplatin history

After 120 years of cisplatin synthesis by Peyrone in 1845 [10], its biological action was

discovered serendipitously during an experiment designed to study the electric current

effect on the growth of Escherichia coli bacteria cell. It was observed that a cell subjected

to electric current formed long filaments but at the cost of reduction in their number, thus

tend to the conclusion that only cell division was repressed but not cell growth. However,

further investigations unveiled that the division stops due to platinum dissolution from

electrode into the medium rather than electric current [11]. Several platinum complexes

tested for anti-proliferative effect on bacteria cells and later on mice revealed cisplatin

has the highest antitumor activity [12]. In 1972, the first clinical trial result was

published [13] and in 1978 it was approved by the FDA (Food and Drug Administration)

for clinical administration [14]. This discovery was a corner stone which persuaded the

interest in anticancer metallo-drugs. Nowadays, cisplatin is used widely as an effective

anticancer drug in the regimen of chemotherapeutic administration, including testicular,

head and neck, ovarian, small cell lung cancer and esophageal cancer [15]. The curing

rate can exceed over 90% in case of testicular and ovarian cancer, if the tumor is

diagnosed in the earlier stage [16, 17]. To improve cisplatin activity by inhibiting

cisplatin-DNA lesions repair, it is also used in combinatorial treatments with other

anticancer drugs like 5-fluorouracil and arabinofuranosylcytosine [18]. Later on, two

analogues drugs, carboplatin and oxaliplatin, received clinical status in United States.

http://en.wikipedia.org/wiki/Antimetabolite#cite_note-1

http://en.wikipedia.org/wiki/Microtubules

http://en.wikipedia.org/wiki/Cell_growth

http://en.wikipedia.org/wiki/Mitosis

http://en.wikipedia.org/wiki/Cancer

http://en.wikipedia.org/wiki/Antigen

http://en.wikipedia.org/wiki/Alemtuzumab

http://en.wikipedia.org/wiki/Bevacizumab

http://en.wikipedia.org/wiki/Trastuzumab

3

Three others, heptaplatin, lobaplatin, and nedaplatin, are extensively used in Asia (Fig

1.1) [19]. These success stories on six platinum-based drugs inspired generations of

bioinorganic chemists and hence far-reaching work has been done in the area of inorganic

chemistry in medicine, witnessed by recently published books and reviews [20].

Figure 1.1: Structures of globally used platinum anticancer drugs.

1.2 Cisplatin mode of action

FDA approved six platinum based drugs (Fig. 1.1) kill cancer cell by complex-DNA

adduct formation [21]. The efficacy of platinum drugs is influenced by how they

internalized by the cell and then nucleus where the critical target, DNA, resides. Initially,

it was thought that passive transport play a significant role in cisplatin cellular uptake,

however, recent studies strongly suggest that active transport through copper transporters

CTR1(Copper TRansport protein 1), CTR2 (Copper TRansport protein 2) and OCT1-3

regulates its routes into the cell [22]. Once inside the cell, the diminished chloride ion

concentration in cytoplasm (2-10 mM) compared to blood (0.1 M) promotes cisplatin

aquation by the subsequent substitution of two chlorides resulting in monoaqua

[Pt(NH3)2Cl(H2O)]+ and diaqua [Pt(NH3)2(H2O)2]

2+ species (Fig. 1.2) [23]. The reactive

electrophilic cationic aquated species that originate from two consecutive aquation steps

of rate constants k1 = 5.18 × 10-5

(t1/2 = 3.4 h) and k2 is 2.75×10-5

(t1/2 = 7 h) [24] form

adducts with protein, RNA and DNA on nucleophilic sites present on these

macromolecules. The nucleophilic attack on Pt(II) square plane occurs through free z-

axis to form five coordinated intermediate. The substitution pattern and nucleophilic

4

preference is governed by trans effect and SHAB concept, respectively. The high

reactivity towards sulfur containing off-target molecules is responsible for the clinical

side effects of cisplatin [25].

Figure 1.2: Different pathways of cisplatin before and after it enters the cell [26].

1.3 Cisplatin toxicity

The anxious side effects associated with platinum chemotherapy are nephrotoxicity,

neurotoxicity, ototoxicity, gastrointestinal toxicity; electrolytic disturbance, fatigue and

decrease in sense of taste, of them the first three are commonly observed.

Nephrotoxicity is a well-known side-effect of cisplatin, which exist mainly in the

proximal tubule part of the kidney, in 20-30% of cancer patients. It is manifested

biochemically by higher creatinine level, lower magnesium and potassium level and

decrease in filtration by glomerular [27]. Cisplatin nephrotoxicity is expressed in

5

different form like acute hypomagnesaemia, distal renal tubular acidosis, renal salts

wasting, hypocalcaemia, hyperuricemia and acute renal failure, for which mechanism is

presented in Fig: 1.3 [28] but the most common is acute kidney injury (AKI) [27].

Kidney tubules are formed by epithelial cells, one of the four basic tissue types present in

the body, of the function to separate interior of the body and various body organs from

the external world, transport of water and mineral salts. Epithelial cells, of distant

structure to do transportation and protection, form cavities of the body, glands, intestine,

urine vessels and kidney tubules. These cells are interconnected through tight junction to

ensure materials passage through epithelial layer and not through the intercellular space.

The concentrations of cisplatin in the proximal tubular epithelial cells are approximately

five times greater than plasma concentrations. Inside renal epithelial cells, cisplatin has

been described to restrict in cytosol, mitochondria, nuclei and lysosomes. As mentioned

earlier, the generated cationic aquated cisplatin, due to low chloride concentration inside

the cell, interact and degrade the lysosomal proteins. Therefore, it has been proposed that

the toxicity may result because of the failure in lysosomal acidification, adversely

affecting its activity [29]. Other factors responsible for renal toxicity are DNA damage,

oxidative stress and inflammation [28].

Figure 1.3: Pathophysiological events in cisplatin nephrotoxicity [30].

6

Neurotoxicity, first reported in 1970, is another dose-limiting side-effect of the cisplatin.

The cisplatin causes neurotoxic effects upon the peripheral nervous system (PNS) and the

central nervous system (CNS), initially characterized by painful paresthesias occurs

during the first few cycles of the drug. After several treatments with cisplatin vibration

sense loss , ataxia and paraesthesia become evident [31]. The degree of platinum-DNA

cross-links in dorsal root ganglion (DRG) neurons at a given additive dose is directly

related with the degree of neurotoxicity. The platinum drugs affect the axons, myelin

sheath, neuronal cell body and the glial structures of the neurons in PNS and CNS.

Although detailed mechanism of neurotoxicity is not yet known but it is suggested that

failure of DNA repair system in neuron cell is responsible for neuronal apoptosis.

Neurotoxicity, due to its irreversible nature, is one of the dangerous cisplatin related side

effects [32].

Ototoxicity is also a commonly encountered side-effect of cisplatin treatment evidenced

by irreversible hearing loss, vertigo and tinnitus. It mainly target three main tissue of

cochlea (inner ear): Corti, spiral ganglion cells and lateral wall (Fig. 1.4). The

accumulation of reactive oxygen species (ROS) like hydroxyl radicals and superoxide

ions are the main causes of ototoxicity, which inhibit antioxidant enzymes resulting in

cisplatin ototoxicity [33].

Figure 1.4: Affected parts of cochlea during ototoxicity.

1.4 Resistance to cisplatin

Despite of consistent rate of initial responses, resistance towards cisplatin is a major

impediment in its successful use, leading to therapeutic failure. Resistance mechanisms

7

arise as a consequence of intracellular changes, which are generally divided into: pre-

binding and post-binding mechanisms [34]. Pre-binding mechanism hinders cisplatin

access to DNA and post-binding mechanism hamper apoptosis, an ultimate result of the

cisplatin DNA interaction.

1.4.1 Pre-binding resistance

There are two mechanisms by which cancer cell evade the cisplatin cytotoxicity prior

reaching the nuclear DNA and other cytoplasmic targets i.e (i) reduced cisplatin

accumulation due to less uptake and greater efflux, and (ii) Inactivation by glutathione

(GSH), methionine, metallothioneins (MT) and other cytoplasmic organelles baring

nucleophilic sites.

1.4.1.1 Decrease cisplatin accumulation

About 20–70% reduction in cisplatin concentration, either due to less uptake or greater

efflux, has been perceived in cisplatin resistant cell lines [35]. It has long been accepted

that cisplatin is internalized by the cell through passive transport but later studies showed

that cisplatin do so through active transport by Cu transporter hCTR1 [36, 37]. It has

been established that copper homeostasis play a significant role in cisplatin uptake.

Reduced expression of hCtr1 protein in several cisplatin resistant cell lines was correlated

with the cisplatin sensitivity levels [38]. Moreover, it was noticed that many cell lines

having resistance to Pt-drugs also display cross-resistance to copper and vice versa [39].

Based upon these results, it has been proposed that Cu and Pt share same transporter

system. It has been noticed that by comparing the resistant and sensitive SR2 cell line of

small cell lung carcinoma, the resistant cell down-regulates the protein expression of

hCtr1 more than half of its corresponding sensitive cell line [40]. Furthermore, the hCtr1

mouse embryonic fibroblasts provide 3-2 fold increase in resistance than their wild-type

transfected cell due to considerably less accumulation of the cisplatin [41]. According to

structural and functional analyses of hCtr1, it consist of 190 amino acids [42] having

extracellular N-terminal and intracellular C-terminal (Fig. 1.5). The C-terminus series

contains His (Histidine) and Cys (Cysteine) residues while N- terminus region has two

His and two Met (Methionine) motifs. Cross intervention of copper and Pt-drugs with

each other by hCtr1 transporter in cell lines indicates that motifs involved in copper

transport will also be important for Pt-drugs. So it has been noted that by the His or Cys

8

motif of C-terminal deletion do not show any effect on Pt-drugs transport, however,

removal of Met-rich motifs from N-terminal of hCtr1 significantly disturb the

transportation, indicating that this motif is involved in cisplatin cross-linkage [43]. In

some cell Na+/K

+ATPase was noted to involve in the cellular uptake of cisplatin. For

instance, in human ovarian cancer cell line the cisplatin accumulation is reduced to 50%

by inhibiting Na+/K

+ATPase via pre-treatment of cells with oubain (inhibitor of Na/K

ATPase), thus indicated seminal role of the cell membrane potential in cisplatin uptake

[44,45].

Figure 1.5: Schematic diagram of hCtr1 structure, showing 190 amino acid residues,

each represented by a circle. The extracellularly located N-terminus, and

intracellularly located C-terminus are indicated. The N-terminal region

contains two each His-rich (yellow) and Met-rich (red) motifs. The C-

terminal His/Cys residues are also indicated [43].

Moreover, benzaldehyde and other similar compounds interrupt intracellular

accumulation of cisplatin presumably due to formation of Schiff bases with integral

membrane transport proteins [46-49].

In addition to the aforesaid mechanisms, reduced cisplatin cellular accumulation might

also occur by higher drug efflux. The multi-drug resistance protein (MRP), a sub class of

ATP-binding cassette (ABC) transporter, has been believed to involve in the Pt-drugs

ejection from the cell [50]. Among seven members (MRP1-7) of MRP, MRP2 (Multidrug

9

resistance-associated protein 2) also known as canalicular multispecific organic anion

transporter 1 (cMOAT) or ATP-binding cassette sub-family C member 2 (ABCC2) play

an imperative role in cisplatin efflux, if bonded with glutathione [51]. After the discovery

of hCTR1 role in cisplatin uptake, interest was diverted towards two Cu transporting P-

type ATPase, ATP7A and ATP7B, which are responsible for extruding Cu from the cell

[52]. These proteins, located in Golgi body apparatus, maintain Cu homeostasis by

forming Cu vesicle and expel them out of the cell membrane. ATP7A is mainly

expressed in small intestine where Cu is absorbed from the nutrient, whereas ATP7B,

mostly present in liver and kidney cells, remove excess of Cu into the bile [53]. In

human, the over expression of ATP7B (epidermoid carcinoma cells) and ATP7A {in

esophageal squamous cell cancer (ESCC)} proteins results in enhancement in cisplatin

resistance [54-55] suggest their involvement in the Pt-drugs efflux (Fig. 1.6).

Figure 1.6: Schematic diagram showing transport of copper and cisplatin in and out of

the cell.

10

1.4.1.2 Inactivation by thiol containing molecules

In addition to platinum-DNA contacts, aquated cisplatin is also diverted to off-target

biological nucleophiles present in the cytoplasm such as GSH, methionine,

metallothioneins and other cysteine-rich proteins, a significant factor contributing in the

drug resistance. These binding events significantly hamper cisplatin access to DNA as

platinum-thiol adducts can be easily kicked out of the cell by MRP2. Furthermore, it

reduces the cytotoxicity by impeding the conversion of mono-adducts to cross-links [56].

The role of off-target platinum-protein interactions is not yet fully explored [57].

1.4.2 Post-binding resistance

For induction of apoptosis (programmed cell death), formation and persistence of

cisplatin-DNA adducts are desirable, however, these intra and inter-strand adducts are

impaired often, and consequently, the resistance cells get hold of repairing or acquire

ability to tolerate the unpaired DNA lesion. Out of the five recognized DNA repair

pathways namely nucleotide excision repair (NER), mismatch repair (MMR), double-

strand break repair, direct repair and base excision repair (BER), the first two

mechanisms play a key role in mediating resistance [58]. NER, a complex biochemical

process, is regulated by several proteins, especially the excision repair cross-

complementation group 1 (ERCC1) protein that play pivotal role in DNA lesion repair

[59, 60]. Elevated level of ERCC1 in the resistance cell line, as revealed by both in vitro

assays and clinical studies, is responsible for preferential removal of lesion and enabling

the DNA repair [62, 62]. The hypersensitivity of testicular cancer cells to cisplatin, more

than 90% success rate, is due to low expression of ERCC1 [63]. Alternatively, other

proteins such as high mobility group (HMG)-domain proteins, for example, potentiate

cisplatin activity by selectively bind to cytotoxic platinum lesions and shielding those

from NER proteins, a phenomenon commonly called as “repair shielding” [64-72].

Hence, high expression of HMG-domain proteins was correlated with hypersensitivity of

the cell to cisplatin [73,74].

In addition, MMR pathway detects and corrects the mismatches before replication and

transcription. MMR consist of number of proteins among those MLH1 and MSH2 are

key for GpG error recognition, and hence less expressed in the resistant cell [75-77].

Apoptosis can be triggered, if cellular damage is surpassed a certain threshold level,

11

nevertheless the value varies from cell to cell. The p35 tumor suppressor gene repair

DNA damage caused by cisplatin, if mutation level is low and promotes apoptosis

otherwise. The deficiency of MMR and dysfunction of p35 tumor suppressor gene enable

the DNA to tolerate the damage [35].

1.5 Monofunctional platinum(II) complexes

Design and development of structurally dissimilar complexes to cisplatin and analogues

is one of many approaches being used to mitigate the damaging side effects (toxicity and

resistance) in Pt-based chemotherapy. In this context, monofunctional platinum(II)

complexes that violate the classical structure-activity relationships (SARs) attracted

special attraction. Much work has been done on monofunctional complexes generally

containing a labile chloride and three non-labile nitrogen-donor ligands (Fig. 1.7). The

term monofunctional refers to its ability to form mono platinum-DNA adduct [78, 79].

Figure 1.7: General structure of monofunctional platinum(II) complexes.

1.5.1 Development of monofunctional platinum complexes

Traditional structure-activity relationships (SARs) established after extensive work on

cisplatin and variants stated that the anticancer activity is administered by charge

neutrality and a square planar L2PtX2 core, where X2 is a pair of cis positioned labile

ligands and L2 a pair of inert ligands in the remaining two sites [80]. Lack of activity,

both in vitro and in vivo, for cationic monofuntional complexes such as [Pt(NH3)3Cl]+

and [Pt(dien)Cl]+ is consistent with the SARs [81]. Despite failure of these particular

complexes, the search for structurally dissimilar complexes to cisplatin was spurred by

inactivity of the latter against all types of cancers and inherent or acquired drug resistance

[82].

Lippard’s research group studied the anticancer activity and mechanism of action of the

cation monofuntional complexes contained two ammine (NH3), a labile ligand and

exocyclic or endocyclic nitrogen-donor heterocyclic ligands. At that time, it was well

Pt

N

NN

Cl (Labile ligand)(Inert ligand)

(Inert ligand) (Inert ligand)

12

accepted that cisplatin exerts anticancer action via bifunctional adduct formation with the

DNA. However, to enhance anticancer efficacy and to curtail side effects of platinum

chemotherapy, cisplatin was used in combination with other anticancer drugs including

DNA intercalators such as doxorubicin. In order to explore the synergistic effect of

platinum compounds and intercalators, Lippard et al made an attempt to platinate the

DNA containing classical intercalator ethidium. An alteration in platination pattern on

the duplex was observed [83, 84] that was attributed to switch in DNA binding mode

from bifunctional to monofunctional, and this proposition was confirmed experimentally

by the subsequent work [85]. In this context, the initial work demonstrated that ethidium

removal on extensive dialysis and inertness of cisplatin to ethidium in solution indicated

lack of ethidium-Pt bond formation. However, succeeding experiments turned down the

idea of no Pt-ethidium covalent bond formation. The term ternary complex composed of

ethidium-Pt-DNA was proposed for the first time by Leng research group [86]. Using

three intercalators, acridine, ethidium and proflavin, they observed that the latter two

tightly bound to DNA and hence could not be removed by extraction, filtration at acidic

pH, or thin-layer chromatography at basic pH. This led to the proposition of ternary

complex formation composed of cis-[Pt(NH3)2]2+

unit, intercalator and DNA. The

proposition was confirmed by florescent measurements, which revealed that the optical

absorption of cis-[Pt(NH3)2(ethidium)Cl]2+

match those of DNA platinated in the

presence of ethidium. Further, it was noted that the DNA promote ligand substitution of

chloride for ethidium [87]. In order to confirm coordination of either of the two exocyclic

nitrogens, N3 and N8, the differential thermochromic behavior was carried out that

uncovered the formation of the N8 regioisomer (Fig. 1.8).

Figure 1.8: Structure of ethidium-containing monofunctional complex.

Pt

H3N NH3

Cl NH2

N

NH2

1

2+

13

A series of cationic monofunctional complexes (Fig. 1.9) carrying a labile chloride of the

general type cis-[Pt(NH3)2(Am)Cl]+, where Am is a nitrogen-donor ligand derived from

pyridine, purine, pyrimidine or aniline, were prepared and screened against in vivo

murine tumor models including S180a, P388, and L1210 [88]. Despite violating the

classical SARs, complexes (2-12) (Fig. 1.10) achieved %ILS greater than that of cisplatin

with the lowest optimum dose (40 mg/kg) was noted for 2 and 7 in the S180a screen.

Complexes 6, 8, 12 and 13 were found active in the L1210 screen and 2, 3 and 6 against

the P388 cell [88].

Figure 1.9: General structure of cationic monofunctional complexes active in the S180a,

P388 and L1210 screen.

The SARs revealed inactive nature of chelating diamine complexes (14, 15) and trans-

amine complexes (16, 17). No effect of the outer sphere chloride or nitrate on activity

indicated that only cationic part play its role in anticancer action. Furthermore, activity

showed reliance on the nature of nitrogen-donor (Am) ligand i.e no activity for

complexes containing primary amine ligands (18-20) and high activity for others

containing heterocyclic secondary or tertiary (Am) ligands. Similarly, activity was

influenced by the nature of leaving group and was markedly changed when chloride (7)

was replaced with bromide (21), most probably due to strong Br-Pt bond (soft-soft

combination). Interestingly, active nature of cis-[Pt(NH3)2(Am)Cl]+ complexes than their

analogous inactive ones (22 and 23) initially tempted Hollis et al to speculate that the

activity stem from bifunctional adduct formation with DNA by ammonia loss. If do so

then ammonia loss should occur at the site trans to chloride to form trans-diammine

complexes, a configuration devoid of activity [88]. Furthermore, lack of correlation

between activity and donor strength of Am also breach the idea of ammonia loss from the

position cis to chloride. This led to the proposition that cis-[Pt(NH3)2(Am)Cl]+ complexes

14

form disruptive and non-repairable monofuntional lesion on DNA stabilized by Am-DNA

interaction. The hypothesis of monofunctional adduct formation was proved

experimentally by two independent research groups; they concluded that platinum-

triamine complexes inhibit DNA synthesis, specifically blocking DNA polymerases at

platinated guanosine residue [89, 90].

Figure 1.10: Structures of monofunctional complexes 2-21.

15

1.5.2 Highly potent monofunctional platinum complexes

Prior work directed that platinum based sterically hindered bifunctional complexes could

provide highly potent anticancer drugs and complex 22 (Fig. 1.11) showed both in vitro

and in vivo higher anticancer activities compared to 23 [91]. According to current reports,

increase in lipophilicity increases the drug uptake through cell membrane and

additionally, with the active role of human OCT transporter [92]. Also, picoplatin, was

designed to overcome drug resistance via reducing drug deactivation. The goal was

achieved by using bulky groups almost normal (103º) to Pt-N2Cl2 core, shielding the

metal from deactivation by off-target bio-molecules [93]. In these regards platinum

complexes with bulky aromatic groups may be anticipated to undergo slower substitution

reactions with nucleophiles than cisplatin [94].

General observation was that monofunctional platinum complexes with steric hindrance

are comparable or more in anticancer action as compared to the cisplatin. A batch of such

monofunctional complexes with various intert N-8-quinolylamide ligands were

synthesized [95] and the results showed that the complex with N-(tert-butoxycarbonyl)-

L-methionine substituent (25), which is the bulkiest and most lipophilic one, is the most

active anticancer candidate [94]. Additionally, complex 25 was active against many of

the cancer cell lines and the anticancer activity was found to be better than the cisplatin

and analogues [94]. For the authentication, same series of platinum complexes was again

tested for their potency and same results were obtained. Anticancer activity of L-valine

(28) and L-leucine-N-8-quinolylamide (29) substituted platinum complexes with respect

to P-388 leukemia cell line was similar to that of the cisplatin and, on the other hand

higher lipophilic complex 30 was more active than 2 (Fig. 1.10) [96].

Two monofunctional complexes, pyriplatin (2) and phenanthriplatin (30), having a less

and a more hydrophobic entity correspondingly, were reacted with methionine and their

kinetics were determined. The complex with more hydrophobic group i.e.

phenanthriplatin showed very slower reaction rate (~10 times) compared to pyriplatin

bearing less hydrophobic group. It is also noteworthy that phenanthriplatin was found

significantly more potent compared to pyriplatin. The explanation for this higher activity

is that the more hydrophobic complexes are more capable to get into the cells and thus

better cellular accumulation takes place [96]. Furthermore, there seems to be a consensus

16

that the connections developed between aromatic rings of the compounds and DNA,

unrecognizable for the NER mechanism are responsible for the anticancer action [96].

Experiments were executed to recognize the mechanism of action of pyriplatin and

phenanthriplatin through RNA polymerase II inhibition studies. Both theoretical [97] and

experimental [97, 98] studies revealed that the mechanism of action of monofunctional

platinum complexes is different from that of cisplatin i.e. gene transcription is handled

herein, via inhibition of Pol II-mediated transcription by the virtue of bulky ligands [97,

98].

Figure 1.11: Structures of bifunctional (22-24) and monofunctional (25-30) Pt(II)

complexes with increasing hydrophobicity and enhanced anticancer

activity.

1.6 Monofunctional platinum complexes with alternative cell kill mechanisms

The monofunctional complexes 31-33 (Fig. 1.12) make covalent complex-DNA

monoadducts with subsequent unwinding of the DNA double strands and thus leading to

the cell death [99, 100]. In these aspects, Pt-acridine compound (33) possessing high

anticancer activity against NSCLC (non-small cell lung cancer) and distinct DNA

binding has been studied widely [101, 102]. The higher anticancer action of the complex

has been attributed to very quick platination rate causing the DNA damage i.e. about 60

17

folds higher compared to cisplatin [103]. Along with these Pt-DNA monoadducts, the

complex also has the ability of intercalation within the DNA bases via acridine moiety

[104]. Nevertheless, literature suggests that like monofunctional complexes based on

other groups lack the cross-linking with DNA for the apoptosis. On the other hand, few

such platinum complexes have the ability of intercalation [105] while few others don’t

have [106]. DNA unwinding essay revealed that some of the monofunctional platinum

compounds don’t alter the DNA structure, as no evidence could be noticed in the essay,

signifying that they induce the cell death via some “non-apoptotic process”; since it is

believed that apoptosis is mediated by DNA interaction of the platinum complexes. In

literature, non-apoptotic processes like autophagy [107], necrosis [106] and parapoptosis

[108] are present in the view of platinum(II) monofunctional complexes.

Figure 1.12: Monofunctional Pt(II) complexes that form Pt-DNA crosslink to induce

apoptosis of the cancer cells.

1.6.1 Autophagy

Although the history of autophagy is around 40 years old [103] but achieved a significant

attention of the researchers in the biological processes recently. Autophagy is a pro-

survival mechanism, in which organelles are self-metabolized in strained situations like

starvation, hypoxia and chemotherapy etc. [109, 110]. It is generally believed that

autophagy process facilitates acquired resistance to anticancer drugs [109, 111, 112]. In

these directions, inhibition of autophagy genes like BECN1, Atg5, by using 3-

methyladenine/chloroquine to mediate apoptosis upon chemotherapy is also a part of the

18

literature [110, 111]. Conversely, recent observations revealed that autophagy may act as

pro-death initiator and ultimately leading to autophagic cell death (25) [113]. This type of

cell death usually depends upon unique cell morphology. Herein, cytoplasm is found to

be encapsulated by a distinct bi-layer (Table 1.1) [112, 113]. The enzyme p53 is

malignant inhibitor that controls the cell response to the DNA breakdown and induces

apoptosis via triggering signal pathways to facilitate transcription and cell cycle arrest

[114, 115]. The conventional platinum based anticancer drugs may induce apoptosis in

tumor cells having wild-type p53 [115-118]. Conversely, mutant p53 present in many of

the tumor cell kinds is incapable of mediating apoptosis (Fig. 1.13), thus leading to drug

resistance [118] (Fig. 1.14). Autophagy, type II apoptotic process is evidenced in

malignant cells cured with monofunctional platinum(II) compounds (25) [107]. The class

of the platinum complexes inhibited Akt signal (cell life sign) and turned on MAPK/Erk

pathway, thus suppressing the mTOR to stimulate autophagy process (Fig. 1.15) [107]. In

this way, monofunctional platinum complexes destroy the tumor cells at the expense of

autophagy, without considering the p53 situation (Fig. 1.14) [118].

Figure 1.13: The fate of cancer cells when they are induced by stress.

19

In these directions, the monofunctional complexes with autophagic pro-death activating

ability are expected to inhibit programmed cell death resistance in cancer treatment [107,

119, 120].

1.6.2 Necrosis

It is a type III mechanism of the cell demise, recognized by its unique morphology i.e.

different from previously mentioned apoptosis and autophagy (Table 1.1) [122, 123].

Necrosis is initiated by tumor necrosis factor via triggering many signal corridors [124]

by unnecessary oxidative products [122, 124], pathogens and venoms [123, 126]. There

is consensus that necrosis does not take place normally, till both of the apoptosis and

autophagy processes were suppressed simultaneously [127]. Remarkably, there are

evidences for the monofunctional platinum compounds, not to bind with DNA covalently

[94].

Table 1.1: Morphology changes of cells via apoptosis [121], autophagy [112],

necrosis [123, 124], and paraptosis [125] cell death pathway.

Morphology Apoptosis Autophagy Necrosis Paraptosis

Cytoplasmic vacuolations Χ

Chromatin condensation Χ Χ Χ

Nuclear fragmentation Χ Χ Χ

Membrane blebbing Χ Χ

Apoptotic bodies Χ Χ Χ

Conversely, the aromatic moieties in the bulky ligands of the platinum complexes set a

stage for additional H-bonding with DNA and so that the complexes act as intercalators

[94]. Many of the inert ligands like Pt-terpyridine (34) (Fig. 1.16) are shown to exhibit

non-covalent interactions with DNA. Interestingly, mechanism of action of the compound

34 revealed that this complex mediates cell death via necrosis [106].

20

Figure 1.14: The possible cell death pathways for cancer cells bearing wild-type p53 or

mutant p53 when treated with cisplatin and monofunctional Pt(II)

compound.

1.6.3 Paraptosis

It is another type of cell death mechanism, although a less familiar one [127], but still

regarded as an effective cell destruction pathway [125]. Paraptosis is an automated non-

apoptotic cell killing pathway [125, 128], which is recognized by the appearance of large

vacuoles in the cytoplasm and lack of the apoptosis [125, 129]. Paraptosis initiated cell

killing may be activated by IGFIR (insulin growth factor I receptor) [127], suppression of

gene transcription or gene translation [129]. The monofunctional platinum complex (35)

(Fig. 1.16) with very bulky inert group is allegedly mediate cell killing via paraptosis

process. The complex is favorably involved with cytoplasmic vacuoles in spite of the

DNA [108].

21

Figure 1.15: Monofunctional Pt(II) compounds suppress Akt and mTOR pathway to

induce autophagy.

Figure 1.16: Monofunctional Pt compounds that did not interact with DNA and exhibit

an alternative cell death pathway.

Above discussion disclose that the nature of carrier ligand and leaving group in classical

platinum drugs is of prime significance to tune the anticancer properties. For example,

22

bidentate chelating carboxylate ligand render slower reactions to carboplatin and

similarly, chelating 1,2 diaminocyclohexane and oxalate ligands make oxaliplatin more

stable and hence low ototoxic and nephrotoxic [130, 131]. Cisplatin and carboplatin form

bifunctional intra-strand cross links in DNA owing to the same carrier ligand (ammine)

[132] while, oxaliplatin forms both inter- and intra-strand cross links with DNA [133] in

the presence of chelating 1,2-diaminocyclohexane ligand. Furthermore, 1,2-

diaminocyclohexane enables oxaliplatin to make bulkier and more hydrophobic

oxaliplatin-DNA adducts than both cisplatin and carboplatin. These bulkier adducts are

able to inhibit DNA replication more effectively compared to cisplatin and carboplatin

[134]. On the other hand, the damaging side effects (toxicity and resistance) of cisplatin

and analogues have been mitigated by using monofunctional platinum(II) complexes

particularly by pyriplatin and phenanthriplatin, having different mechanism of actions.

This suggests that nature of the attached ligands is also important in tuning the

mechanism of action along with anticancer properties of the monofunctional platinum(II)

complexes i.e. for instance, hydrophobic phenanthridine ligand is found to be responsible

for greater cellular uptake and potency of phenanthriplatin as compared to pyriplatin.

Motivated by these observations we are investigating a new type of neutral

monofunctional platinum(II) complexes having two bulky and hydrophobic inert ligands

(a chelating dithiocarbamate and an organophosphine) and chloride as a labile ligand

(Fig. 1.17).

Figure 1.17: General structure of the complex having two inert ligands and a labile

ligand

Pt

Cl

P S

S

N

X

N R

X

X

(Inert ligand)

(Inert ligand)(Labile ligand)

https://en.wikipedia.org/wiki/Nephrotoxicity

23

These complexes are expected to have lower resistance because of low reactivity towards

off-target sulfur containing bio-molecules due to strong binding of the dithiocarbamate

and organophosphine (SHAB concept) ligands. Lower toxicity may be anticipated for

these complexes, as dithiocarbamates are being used as inhibitors of cisplatin induced

nephrotoxicity and renal toxicity [135]. Likewise, potential shielding of platinum square

plane by aromatic C-H groups of organophosphine and also greater cellular uptake

because of more bulky and hydrophobic ligands make them attractive candidates for the

anticancer investigations. The mechanism of action of these complexes is expected to be

different from that of cisplatin and analogues due to different structural chemistry of the

attached ligands.

24

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Chapter 2

EXPERIMENTAL

38

2.1 Materials

Precursors such as benzylpiperazine, 1-(2-furoyl)piperazine, diphenylmethylpiperazine,

4-nitrophenylpiperazine, 4-methoxyphenylpiperazinne, 4-hydroxyphenylpiperazine, 2-

hydroxyethylpiperazine, 4-(3-(piperidin-3-yl)propyl)piperidine and tris-(4-flourophenyl)-

phosphine were purchased from Sigma-Aldrich. Other precursors such as tris-(4-

chlorophenyl)phosphine, tris-(p-tolyl)phosphine, (diphenyl-p-tolyl)phosphine, bis-

diphenyldiphosphinobutane and platinum(II) chloride were procured from Wako Japan

and carbon disulfide from Riedel-de Haën. All the reagents were used without further

purification owing to their analytical grade quality. Solvents obtained from Daejung,

Sigma-Aldrich and Scharlau were dried-up by standard methods [1, 2]. CT-DNA (calf

thymus), acetic acid, ethylenediaminetetraacetic acid disodium salt, penicillin-G, pyruvic

acid, L-glutamine, sodium dodecyl sulfate (SDS), sodium sarcosinate, sodium chloride,

streptomycin sulfate, thiazolyl blue tetrazolium bromide (MTT), Triton X-100 and trizma

base were also acquired from Sigma-Aldrich. Dulbecco’s Modified Eagle Medium

(DMEM) and fetal bovine serum (FBS) were purchased from GibcoBRL, Gaithersburg,

MD.

2.2 Instrumentation

Melting points were determined in a capillary tube using Gallenkamp (U.K) electro-

thermal melting point apparatus. IR spectra in the range of 4000-200 cm-1

were acquired

on a Thermo Scientific Nicolet-6700 FT-IR Spectrophotometer. Elemental analysis was

performed using a CE-440 Elemental Analyzer (Exeter Analytical, Inc.). 1H,

13C and

31P

NMR were recorded on a Bruker instrument using TMS (1H and

13C) and phosphoric acid

(31

P) as an internal reference. Chemical shifts are presented in ppm and coupling constant

(J) values are presented in Hz. The multiplicities of signals in 1H NMR are given with

chemical shifts (s = singlet, d = doublet, t = triplet, dd = doublet of a doublet, m =

multiplet). The X-ray single crystal analysis of the crystalline complexes was performed

by Bruker Kapa APEX-II CCD diffractometer assembled with a CCD detector set 40.0

mm from crystal. Different intensities were measured from a sealed ceramic diffraction

tube (SIEMENS) having monochromatic Mo-Kα radiation. Structures of the complexes

were solved by using Patterson and DIRDIF method. Further refinement on F2 was

39

performed by full matrix least square technique using SHELXL-97 [3]. The crystals were

kept at 296(2) K during data collection.

2.3 General procedure for synthesis of the ligands

The dithiocarbamate ligands (1-9) were synthesized as sodium salts, by stirring equal

molar solutions of the corresponding precursor (piperazine/piperidine/morpholine based)

and sodium hydroxide in methanol for 1 hour and then slow drop wise addition of equal

molar methanolic solution of CS2, in ice bath conditions with 4 hours stirring (Scheme

2.1). The resulting solutions were filtered off, rota-evaporated. The obtained solid

products were dried and recrystallized in methanol. The ligand 10, containing two

dithiocarbamate functionalities was also prepared using the same method however, molar

ratios for the corresponding precursor, sodium hydroxide and CS2 were 1:2:2,

respectively (Scheme 2.2).

R ZNH

NaOH,+

5h stirring

Dry methanol,R Z

N SNa

S

1C SS

-H2O

H2CHOR =

(L-1)

H3COO

C

O

(L-2) (L-3)

O2N

(L-4) (L-5) (L-6)

HO

(L-7)

Where Z = Nitrogren (L-1-L-7), Oxygen (L-8), Carbon (L-9)

Scheme 2.1: Synthesis of the ligands L-1 to L-9.

2NaOH,+

5h stirring

Dry methanol,C SS

-H2O2(H2C)3 NH

HN

(CH2)3 NN

S

SNa

S

NaS

Scheme 2.2: Synthesis of the ligand L-10.

40

Sodium 4-(4-methoxyphenyl)piperazine-1-carbodithioate (L-1)

1

2

2'

3

3'

N

N

OH3C

SNa

S4

56

5'6'

78

Amount of reagents used: 2 g (10.4 mmol) 1-(4-methoxyphenyl)piperazine, 0.42 g

(10.4 mmol) sodium hydroxide and 0.79 g (10.40 mmol) carbon disulfide. Yield: 2.95 g,

92%. m.p. C. Elemental analysis, % Calculated (Found) for C12H15N2OS2Na (Mol.

Wt: 290.4 g/mol): C, 49.63 (49.60); H, 5.21 (5.20); N, 9.65 (9.63); S, 22.08 (22.07).

FT-IR (4000-400 cm-1

): 1441 ν(N-CSS), 1217 ν(C-NC2), 1018, 1001 ν(C-S).

Sodium 4-(2-furoyl)piperazine-1-carbodithioate (L-2)

2

2'

3

3'

N

N

SNa

S

1

456

7

8

O

O

Amount of reagents used: 1.87 g (10.4 mmol) 1-(2-furoyl)piperazine, 0.42 g (10.4

mmol) sodium hydroxide and 0.79 g (10.40 mmol) carbon disulfide. Yield: 2.93 g, 95%.

m.p. 212-214 C. Elemental analysis, % Calculated (Found) for C10H11N2O2S2Na (Mol.

Wt: 278.3 g/mol): C, 43.15 (43.10); H, 3.98 (3.96); N, 10.06 (10.02); S, 23.04 (23.00).

FT-IR (4000-400 cm-1

): 1601 ν(C=O), 1433 ν(N-CSS), 1209 ν(C-NC2), 1015, 1003 ν(C-

S).

Sodium 4-diphenylmethylpiperazine-1-carbodithioate (L-3)

2

2'

3

3'

N

N

SNa

S

1

456

6'7

7'8

Amount of reagents used: 2.5 g (10 mmol) 1-diphenylmethylpiperazine, 0.4 g (10

mmol) sodium hydroxide and 0.76 g (10 mmol) carbon disulfide. Yield: 3.18 g, 87%.

41

m.p. 229-230 C. Elemental analysis, % Calculated (Found) for C18H19N2NaS2 (Mol. Wt:

350.47 g/mol): C, 61.69 (61.67); H, 5.46 (5.44); N, 7.99 (7.98); S, 18.30 (18.28). FT-IR

(4000-400 cm-1

): 1450 ν(N-CSS), 1209 ν(C-NC2), 1026, 993 ν(C-S).

Sodium 4-(4-nitrophenyl)piperazine-1-carbodithioate (L-4)

2

2'

3

3'

N

N

O2N

SNa

S4

56

5'6'

7

1

Amount of reagents used: 2.07 g (10 mmol) 1-(4-nitrophenyl)piperazine, 0.4 g (10


m.p. 217-220 C. Elemental analysis, % Calculated (Found) for C11H12N3NaO2S2 (Mol.

Wt: 305.35 g/mol): C, 43.27 (43.25); H, 3.96 (3.93); N,13.76 (13.75); S, 21.01 (21.00).

FT-IR (4000-400 cm-1

): 1597, 1308 ν(N=O), 1474 ν(N-CSS), 1211 ν(C-NC2), 1007, 993

ν(C-S).

Sodium 4-(2-hydroxyethyl)piperazine-1-carbodithioate (L-5)

2

2'

3

3'

N

N

SNa

S

1

45HO

Amount of reagents used: 1.95 g (15 mmol) 1-(2-hydroxyethyl)piperazine, 0.6 g (15


m.p. 212-215 C. Elemental analysis, % Calculated (Found) for C7H13N2NaOS2 (Mol.

Wt: 228.31 g/mol): C, 36.82 (36.80); H, 5.74 (5.71); N, 12.27 (12.25); S, 28.09 (28.07).

FT-IR (4000-400 cm-1

): 3343 ν(O-H), 1454 ν( N-CSS), 1215 ν(C-NC2), 1009, 991 ν(C-

S).

Sodium 4-benzylpiperazine-1-carbodithioate (L-6)

2

2'

3

3'

N

N

SNa

S

1

456

6'7

7'8

42

Amount of reagents used: 1.83 g (10.4 mmol) 1-benzylpiperazine, 0.42 g (10.4 mmol)

sodium hydroxide and 0.79 g (10.4 mmol) carbon disulfide. Yield: 2.67 g, 88%. m.p. 220

C. Elemental analysis, % Calculated (Found) for C12H15N2NaS2 (Mol. Wt: 274.4 g/mol):

C, 52.53 (52.51); H, 5.51 (5.50); N, 10.21 (10.20); S, 23.37 (23.35). FT-IR (4000-400

cm-1

): 1466 ν(N-CSS), 1215 ν(C-NC2), 1028, 997 ν(C-S).

Sodium 4-(4-hydroxyphenylyl)piperazine-1-carbodithioate (L-7)

2

2'

3

3'

N

N

HO

SNa

S4

56

5'6'

7

1

Amount of reagents used: 1.78 g (10 mmol) 1-(4-hydroxyphenyl)piperazine, 0.4 g (10


m.p. 222-224 C. Elemental analysis, % Calculated (Found) for C11H13N2NaOS2 (Mol.

Wt: 276.4 g/mol): C, 47.81 (47.77); H, 4.74 (4.71); N,10.14 (10.10); S, 23.21 (23.18).

FT-IR (4000-400 cm-1

): 1454 ν(N-CSS), 1221 ν(C-NC2), 1047, 1003 ν(C-S).

Sodium morpholine-4-carbodithioate (L-8)

NaS

S

NO

12

2'

3

3'

Amount of reagents used: 1 g (11.48 mmol) morpholine, 0.46 g (11.48 mmol) sodium

hydroxide and 0.87 g (11.48 mmol) carbon disulfide. Yield: 2 g, 86%. m.p. 208-212 C.

Elemental analysis, % Calculated (Found) for C5H8NNaOS2 (Mol. Wt: 185.2 g/mol): C,

32.42 (32.39); H, 4.35 (4.30); N, 7.56 (7.51); S, 34.62 (34.59). FT-IR (4000-400 cm-1

):

1463 ν(N-CSS), 1213 ν(C-NC2), 1024, 985 ν(C-S).

Sodium piperidine-1-carbodithioate (L-9)

NaS

S

N1

2

2'3

3'

4

43

Amount of reagents used: 0.862 g (10.12 mmol) piperidine, 0.405 g (10.12 mmol)

sodium hydroxide and 0.77 g (10.4 mmol) carbon disulfide. Yield: 1.73 g, 85%. m.p.

226-228 C. Elemental analysis, % Calculated (Found) for C6H10NNaS2 (Mol. Wt: 183.3

g/mol): C, 39.32 (39.27); H, 5.50 (5.48); N, 7.64(7.60); S, 34.99 (34.96). FT-IR (4000-

400 cm-1

): 1470 ν(N-CSS), 1215 ν(C-NC2), 1020, 999 ν(C-S).

Sodium 4,4ʹ-trimethylenedipiperidine-1-carbodithioate (L-10)

NaS

S

N

SNa

S

N1

2

2'

3

3'

4 5

6

Amount of reagents used: 2.1 g (10 mmol) 4-(3-(piperidin-3-yl)propyl)piperidine, 0.4 g

(10 mmol) sodium hydroxide and 0.76 g (10 mmol) carbon disulfide. Yield: 2.96 g, 91%.

m.p. 204-206 C. Elemental analysis, % Calculated (Found) for C15H24N2Na2S4 (Mol.

Wt: 406.6 g/mol): C, 44.31 (44.30); H, 5.95 (5.92); N, 6.89 (6.87); S, 31.54 (31.52).

FT-IR (4000-400 cm-1

): 1464 ν(N-CSS), 1207 ν(C-NC2), 1026, 999 ν(C-S).

2.4 General procedure for the synthesis of hetroleptic Pt(II) complexes

Heteroleptic platinum(II) complexes (1-37) containing dithiocarbamate, organophosphine

and chloride ligands were synthesized by simultaneous and dropwise addition of the

corresponding organophosphine (1 mmol) in chloroform and sodium salt of the dithio-

carbamate ligand (1 mmol) in methanol into the chloroform suspension of platinum(II)

chloride (1 mmol ) and refluxing the reaction mixture for 5 hours (Schemes 2.3 & 2.4).

The resulting solutions were filtered off and rota-evaporated to get the solid products,

which were then dissolved in acetone to remove sodium chloride, a side product of the

reaction. After filtration, rota-evaporation and drying, the products were then

recrystallized in chloroform and methanol.

Dinuclear heteroleptic platinum(II) complexes (38-42) were prepared adopting the same

general procedure however, molar ratios for the platinum(II) chloride, corresponding

organophosphine and dithiocarbamate ligands were 2:2:1 respectively (Schemes 2.5 &

2.6).

44

PtCl2 + RPR'3 ZN SNa

S

+

MethanolChloroform R Z

N

S

S

Pt

Cl

-NaCl PR'3

R, Z = 4-methoxyphenyl, nitrogen (1-4), R, Z = 1-(2-furoyl), nitrogen (6-9),

R, Z = 1-diphenylmethyl, nitrogen (11-14), R, Z = 4-nitrophenyl, nitrogen (16-18),

R, Z = 2-hydroxyethyl, nitrogen (20-23), R, Z = 1-benzyl, nitrogen (25-27),

R, Z = 4-hydroxyphenyl, nitrogen (29-32), Z = oxygen (34-35), Z = carbon (36-37)

Scheme 2.3: Synthesis of the complexes 1-4, 6-9, 11-14, 16-18, 20-23, 25-27, 29-32 and

34-37 where PRʹ3 = tris-p-flourophenylphosphine (1, 6, 11, 16, 20, 25, 29),

tris-p-chlorophenylphosphine (2, 7, 12, 21, 30), diphenyl-p-tolylphosphine

(3, 8, 13, 17, 22, 26, 31, 34, 36), tri-p-tolylphosphine (4, 9, 14, 18, 23, 27,

32) and tris-p-methoxyphenylphosphine (35, 37).

PtCl2 + RPh2P(CH2)4 PPh2 NN SNa

S

+

MethanolChloroform

-NaCl

Pt

P S

S

NN R

P

Cl

Scheme 2.4: Synthesis of the complexes 5, 10, 15, 19, 24, 28 and 33, where R = 4-

methoxyphenyl (5), 1-(2-furoyl) (10), 1-diphenylmethyl (15), 4-nitrophenyl

(19), 2-hydroxyethyl (24), 1-benzyl (28), and 4-hydroxyphenyl (33).

45

NaS

S

N(CH2)3

SNa

S

N

Pt

Cl

P S

S

R'N

R'R'

(CH2)3Pt

Cl

PS

S

R'N

R' R'

2 PtCl2 + 2 PR'3 +

MethanolChloroform-2 NaCl

Scheme 2.5: Synthesis of the complexes 38-41, where PRʹ3 = tris-p-flouro-

phenylphosphine (38), tris-p-chlorophenylphosphine (39), diphenyl-p-

tolylphosphine (40) and tri-p-tolylphosphine (41).

Pt

P S

S

N(CH2)3

P

Cl2Pt

PS

S P

N

2 PtCl2 + 2 Ph2P(CH2)4 PPh2 +

NaS

S

N(CH2)3

SNa

S

N

MethanolChloroform-2 NaCl

Scheme 2.6: Synthesis of the complex 42.

The complexes 29a, 29b, 29c, 32a, 32b, 34a, 34b and 34c were prepared from 29, 32 and

34 correspondingly, by replacing the chloride. In procedure, the respective complex (1

mmol) was reacted with XY reagent (1 mmol) in a mixture of acetonitrile and methanol

46

followed by 3 hours stirring at room temperature. After the consecutive processes of

filtration and rota-evaporation, a solid product was obtained (Scheme 2.7).

R ZN

S

S

Pt

Cl

PR'3

+ XYstirring 3h

-XClR Z

N

S

S

Pt

Y

PR'3

Scheme 2.7: Synthesis of the complexes 29a, 29b, 29c, 32a, 32b, 34a, 34b and 34c by

chloride exchange reactions, where Z = oxygen (29a, 29b, 29c), R = 4-

hydroxyphenyl and Z = nitrogen (32a, 32b, 34a, 34b and 34c), XY =

NaSCN (29a, 32a, 34a), NH4Br (29b, 32b, 34b), KI (29c, 34c), PRʹ3 =

diphenyl-p-tolylphosphine (29a, 29b, 29c), tris-p-flourophenylphosphine

(34a, 34b, 34c), tri-p-tolylphosphine (32a, 32b).

Chlorido[4-(4-methoxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophe-

nylphosphine)]platinum(II) (1)

Pt

Cl

P S

S

N

F

N OCH312

3

3'

2'

4

5 6

7

a

b

cd

F

F

6'5'

8

Amount of reagents used: 0.29 g (1 mmol) sodium 4-(4-methoxyphenyl)piperazine-1-

carbodithioate, 0.266 g (1 mmol) platinum(II) chloride and 0.316 g (1 mmol) tris-p-

flourophenylphosphine. Yield: 0.66 g, 82%. m.p. C. Elemental analysis, %

Calculated (Found) for C30H27ClF3N2OPPtS2 (Mol. Wt: 814.2 g/mol): C, 44.26 (44.24);

H, 3.34 (3.33); N, 3.44 (3.42); S, 7.88 (7.86). FT-IR (4000-200 cm-1

): 1495 ν(N-CSS),

1225 ν(C-NC2), 1013 ν(C-S), 359 ν(Pt-S), 299 ν(Pt-Cl), 239 ν(Pt-P).

Chlorido[4-(4-methoxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-chlorophe-


47

Pt

Cl

P S

S

N

Cl

N OCH31

2

3

3'

2'

4

5 6

7

a

b

cd

Cl

Cl

6'5'

8



chlorophenylphosphine. Yield: 0.69 g, 81%. m.p. C. Elemental analysis, %

Calculated (Found) for C30H27Cl4N2OPPtS2 (Mol. Wt: 863.5 g/mol): C, 41.73 (41.71); H,

3.15 (3.12); N, 3.24 (3.21); S, 7.43 (7.41). FT-IR (4000-200 cm-1

): 1479 ν(N-CSS), 1238

ν(C-NC2), 1011 ν(C-S), 361 ν(Pt-S), 293 ν(Pt-Cl), 239 ν(Pt-P).

Chlorido[4-(4-methoxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolyl

phosphine)]platinum(II) (3)

Pt

Cl

P S

S

NN OCH3

12

3

3'

2'

4

5 6

7

a

b

cd

6'5'

8


carbodithioate, 0.266 g (1 mmol) platinum(II) chloride and 0.276 g (1 mmol) diphenyl-p-

tolylphosphine. Yield: 0.61 g, 79%. m.p. 245 C. Elemental analysis, % Calculated

(Found) for C31H32ClN2OPPtS2 (Mol. Wt: 774.2 g/mol): C, 48.09 (48.06); H, 4.17 (4.13);

N, 3.62 (3.60); S, 8.28 (8.26). FT-IR (4000-200 cm-1

): 1508 ν(N-CSS), 1227 ν(C-NC2),

1016 ν(C-S), 377 ν(Pt-S), 279 ν(Pt-Cl), 227 ν(Pt-P).

Chlorido[4-(4-methoxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphos-

phine)]platinum(II) (4)

Pt

Cl

P S

S

NN OCH3

12

3

3'

2'

4

5 6

7

a

b

cd

6'5'

8

48



tolylphosphine. Yield: 0.61 g, 76%. m.p. 238-241 C. Elemental analysis, % Calculated


N, 3.49 (3.48); S, 7.99 (7.96). FT-IR (4000-200 cm-1

): 1510 ν(N-CSS), 1227 ν(C-NC2),


4-(4-methoxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis(diphenylphosphino)

butane)platinum(II) chloride (5)

Pt

P S

S

NN

P

ClOCH312

3

2'

3'

4

5 6

7

6'5'

a

b

cd

ef

8


carbodithioate, 0.266 g (1 mmol) platinum(II) chloride and 0.426 g (1 mmol) 1,4-bis

(diphenylphosphino)butane. Yield: 0.66 g, 76%. m.p. 223-226 C. Elemental analysis, %

Calculated (Found) for C40H43ClN2OP2PtS2 (Mol. Wt: 924.4 g/mol): C, 51.97 (51.95); H,

4.69 (4.65); N, 3.03 (3.00); S, 6.94 (6.92). FT-IR (4000-200 cm-1

): 1504 ν(N-CSS), 1233

ν(C-NC2), 1015, 1002 ν(C-S), 372 ν(Pt-S), 240 ν(Pt-P).

Chlorido-4-(2-furoyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophenylphos-

phine)platinum(II) (6)

Pt

Cl

P S

S

N

F

N

O

O1

2

3

2'

3'

4 5 6

7

8

a

b

cd

F

F

Amount of reagents used: 0.278 g (1 mmol) sodium 4-(2-furoyl)piperazine-1-


flourophenylphosphine. Yield: 0.66 g, 82%. m.p. C. Elemental analysis, %

Calculated (Found) for C28H23ClF3N2O2PPtS2 (Mol. Wt: 802.1 g/mol): C, 41.93 (41.91);

49

H, 2.89 (2.88); N, 3.49 (3.47); S, 8.00 (7.98). FT-IR (4000-200 cm-1

): 1636 ν(C=O),

1493 ν(N-CSS), 1223 ν(C-NC2), 1009 ν(C-S), 361 ν(Pt-S), 287 ν(Pt-Cl), 220 ν(Pt-P).

Chlorido-4-(2-furoyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-chlorophenylphos-


Pt

Cl

P S

S

N

Cl

N

O

O1

2

3

2'

3'

4 5 6

7

8

a

b

cd

Cl

Cl



chlorophenylphosphine. Yield: 0.62 g, 73%. m.p. 2 C. Elemental analysis, %

Calculated (Found) for C28H23Cl4N2O2PPtS2 (Mol. Wt: 851.5 g/mol): C, 39.50 (39.47);

H, 2.72 (2.70); N, 3.29 (3.26); S, 7.53 (7.52). FT-IR (4000-200 cm-1

): 1626 ν(C=O),

1479 ν(N-CSS), 1241 ν(C-NC2), 1011 ν(C-S), 378 ν(Pt-S), 297 ν(Pt-Cl), 220 ν(Pt-P).

Chlorido-4-(2-furoyl)piperazine-1-carbodithioato-κ2S,Sʹ(diphenyl-p-tolylphosphine)

platinum(II) (8)

Pt

Cl

P S

S

NN

O

O1

2

3

2'

3'

4 5 6

7

8

a

b

cd




(Found) for C29H28ClN2O2PPtS2 (Mol. Wt: 762.2 g/mol): C, 45.70 (45.69); H, 3.70

(3.68); N, 3.68 (3.65); S, 8.41 (8.40). FT-IR (4000-200 cm-1

): 1620 ν(C=O), 1479 ν(N-

CSS), 1242 ν(C-NC2), 999 ν(C-S), 366 ν(Pt-S), 297 ν(Pt-Cl), 235 ν(Pt-P).

Chlorido-4-(2-furoyl)piperazine-1-carbodithioato-κ2S,Sʹ(tris-p-tolylphosphine)plati-

num(II) (9)

50

Pt

Cl

P S

S

NN

O

O1

2

3

2'

3'

4 56

7

8

a

b

cd





(4.05); N, 3.54 (3.53); S, 8.12 (8.10). FT-IR (4000-200 cm-1

): 1620 ν(C=O), 1499 ν(N-

CSS), 1242 ν(C-NC2), 1001 ν(C-S), 368 ν(Pt-S), 278 ν(Pt-Cl), 212 ν(Pt-P).

4-(2-furoyl)piperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis(diphenylphosphino)butane)-

platinum(II) chloride (10)

Pt

P S

S

NN

P

Cl1

2

3

2'

3'

4 5 6

7

a

b

cd

ef

8

O

O



(diphenylphosphino)butane. Yield: 0.59 g, 69%. m.p. 220 C. Elemental analysis, %

Calculated (Found) for C38H39ClN2O2P2PtS2 (Mol. Wt: 912.3 g/mol): C, 50.03 (50.00); H,

4.31 (4.29); N, 3.07 (3.05); S, 7.03 (7.00). FT-IR (4000-200 cm-1

): 1622 ν(C=O), 1503

ν(N-CSS), 1241 ν(C-NC2), 1013, 996 ν(C-S), 359 ν(Pt-S), 227 ν(Pt-P).

Chlorido(4-diphenylmethylpiperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophenyl-

phosphine)platinum(II) (11)

51

Pt

Cl

P S

S

NF

FF

N1

2

3

4

5 6

7

87'

6'

3'

2'

a

b

cd

Amount of reagents used: 0.35 g (1 mmol) sodium 4-diphenylmethylpiperazine-1-


flourophenylphosphine. Yield: 0.62 g, 71%. m.p. 2 C. Elemental analysis, %

Calculated (Found) for C36H31ClF3N2PPtS2 (Mol. Wt: 874.3 g/mol): C, 49.46 (49.42); H,

3.57 (3.54); N, 3.20 (3.17); S, 7.34 (7.31). FT-IR (4000-200 cm-1

): 1495 ν(N-CSS), 1229


Chlorido(4-diphenylmethylpiperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-chlorophenyl-

phosphine)platinum(II) (12)

Pt

Cl

P S

S

NCl

ClCl

N1

2

34

5 6

7

87'

6'

3'

2'

a

b

cd




Calculated (Found) for C36H31Cl4N2PPtS2 (Mol. Wt: 923.6 g/mol): C, 46.81(46.80); H,

3.38 (3.35); N, 3.03 (3.01); S, 6.94 (6.92): FT-IR (4000-200 cm-1

): 1479 ν(N-CSS), 1244


Chlorido(4-diphenylmethylpiperazine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylpho-

sphine)platinum(II) (13)

Pt

Cl

P S

S

NN

12

3

4

5 6

7

87'

6'

a

2'

3'

b

cd

52




(Found) for C37H36ClN2PPtS2 (Mol. Wt: 834.3 g/mol): C, 53.26 (53.22); H, 4.35 (4.32);

N, 3.36 (3.33); S, 7.69 (7.66). FT-IR (4000-200 cm-1

): 1520 ν(N-CSS), 1244 ν(C-NC2),


Chlorido(4-diphenylmethylpiperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphosphin-

e)platinum(II) (14)

Pt

Cl

P S

S

NN

12

3

2'

3'4

5 6

7

87'

6'

a

b

cd





N, 3.25 (3.23); S, 7.44 (7.42). FT-IR (4000-200 cm-1

): 1530 ν(N-CSS), 1244 ν(C-NC2),


(4-diphenylmethylpiperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis-(diphenylphosphino)-


Pt

P S

S

NN

P

Cl1

2

3

2'

3'4

5 6

7

87'

6'

ef

a

b

cd




53

Calculated (Found) for C46H47ClN2P2PtS2 (Mol. Wt: 984.5 g/mol): C, 56.12(56.10); H,

4.81 (4.80); N, 2.85 (2.83); S, 6.51(6.50). FT-IR (4000-200 cm-1

): 1530 ν(N-CSS), 1244


Chlorido[4-(4-nitrophenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophenyl-


Pt

Cl

P S

S

N

F

N NO212

3

3'

2'

4

5 6

7

a

b

cd

F

F

6'5'

Amount of reagents used: 0.305 g (1 mmol) sodium 4-(4-nitrophenyl)piperazine-1-



Calculated (Found) for C29H24ClF3N3O2PPtS2 (Mol. Wt: 829.1 g/mol): C, 42.01 (42.00);

H, 2.92 (2.90); N, 5.07 (5.05); S, 7.73 (7.71). FT-IR (4000-200 cm-1

): 1587, 1317

ν(N=O), 1495 ν(N-CSS), 1227 ν(C-NC2), 1011 ν(C-S), 361 ν(Pt-S), 293 ν(Pt-Cl), 233

ν(Pt-P).

Chlorido[4-(4-nitrophenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylpho-

sphine)]platinum(II) (17)

Pt

Cl

P S

S

NN NO2

12

3

3'

2'

4

5 6

7

a

b

cd





(3.68); N, 5.32 (5.30); S, 8.13 (8.10). FT-IR (4000-200 cm-1

): 1591, 1315 ν(N=O), 1508

ν(N-CSS), 1229 ν(C-NC2), 1009 ν(C-S), 366 ν(Pt-S), 299 ν(Pt-Cl), 222 ν(Pt-P).

54

Chlorido[4-(4-nitrophenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphos-


Pt

Cl

P S

S

NN NO2

12

3

3'

2'

4

5 6

7

a

b

cd

6'5'





(4.06); N, 5.14 (5.13); S, 7.85 (7.84). FT-IR (4000-200 cm-1

): 1595, 1323 ν(N=O), 1497


4-(4-nitrophenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis-(diphenylphosphino)-


Pt

P S

S

NN

P

ClNO21

2

3

2'

3'

4

5 6

7

6'5'

ef

a

b

cd



(diphenylphosphino)butane. Yield: 0.64 g, 73%. m.p. 236 C. Elemental analysis, %

Calculated (Found) for C39H40ClN3O2P2PtS2 (Mol. Wt: 939.4 g/mol): C, 49.87 (49.84);

H, 4.29 (4.26); N, 4.47 (4.44); S, 6.83 (6.81). FT-IR (4000-200 cm-1

): 1589, 1310

ν(N=O), 1504 ν(N-CSS), 1229 ν(C-NC2), 1009, 997 ν(C-S), 372 ν(Pt-S), 222 ν(Pt-P).

Chlorido[4-(2-hydroxyethyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophenyl-


55

Pt

Cl

P S

S

NF

FF

NOH1

2

3

3'

2'

4

5

a

b

cd

Amount of reagents used: 0.228 g (1 mmol) sodium 4-(2-hydroxyethyl)piperazine-1-


flourophenylphosphine. Yield: 0.59 g, 78%. m.p. 235-237 C. Elemental analysis, %


H, 3.35 (3.32); N, 3.72 (3.71); S, 8.53 (8.51). FT-IR (4000-200 cm-1

): 3306 ν(O-H), 1495


Chlorido[4-(2-hydroxyethyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-chlorophe-


Pt

Cl

P S

S

NCl

ClCl

NOH1

2

3

3'

2'

4

5

a

b

cd



chlorophenylphosphine. Yield: 0.6 g, 75%. m.p. 228-230 C. Elemental analysis, %


3.14 (3.10); N, 3.50 (3.46); S, 8.00 (7.98). FT-IR (4000-200 cm-1

): 3337 ν(O-H), 1479


Chlorido[4-(2-hydroxyethyl)piperazine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylph-

osphine)]platinum(II) (22)

Pt

Cl

P S

S

NN

OH12

3

3'

2'

4

5

a

b

cd

56





N, 3.93 (3.90); S, 9.00 (8.97). FT-IR (4000-200 cm-1

): 3329 ν(O-H), 1529 ν(N-CSS),


Chlorido[4-(2-hydroxyethyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphos-


Pt

Cl

P S

S

NN

OH12

3

3'

2'

4

5

a

b

cd





N, 3.78 (3.77); S, 8.66 (8.63). FT-IR (4000-200 cm-1

): 3325 ν(O-H), 1526 ν(N-CSS),


4-(2-hydroxyethyl)piperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis-(diphenylphosphino)-


Pt

P S

S

NN

P

ClOH1

2

3

2'

3'

4

5

a

b

cd

ef




57


4.79 (4.76); N, 3.25 (3.22); S, 7.44 (7.41). FT-IR (4000-200 cm-1

): 3316 ν(O-H), 1528

ν(N-CSS), 1252 ν(C-NC2), 1030, 989 ν(C-S), 361 ν(Pt-S), 212 ν(Pt-P).

Chlorido(4-benzylpiperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophenylphos-


PtCl

PS

S

NF

FF

N1

2

3

4

5

6

7 8

6'

7'

2'

3'

a

b

cd

Amount of reagents used: 0.274 g (1 mmol) sodium 4-benzylpiperazine-1-



Calculated (Found) for C30H27ClF3N2PPtS2 (Mol. Wt: 798.2 g/mol): C, 45.14 (45.10); H,

3.41 (3.38); N, 3.51 (3.47); S, 8.03 (8.00). FT-IR (4000-200 cm-1

): 1495 ν(N-CSS), 1227


Chlorido(4-benzylpiperazine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylphosphine)-

platinum(II) (26)

PtCl

P S

S

NN

12

2'

3

3'4

5

6

7 8

7'

6'

a

b

cd





N, 3.69 (3.67); S, 8.46 (8.45). FT-IR (4000-200 cm-1

): 1497 ν(N-CSS), 1238 ν(C-NC2),


58

Chlorido(4-benzylpiperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphosphine)-

platinum(II) (27)

PtCl

P S

S

NN

12

2'

3

3'4

5

6

7 8

7'

6'

a

b

cd





N, 3.56 (3.54); S, 8.16 (8.15). FT-IR (4000-200 cm-1

): 1531 ν(N-CSS), 1240 ν(C-NC2),


(4-benzylpiperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis-(diphenylphosphino)butane)

platinum(II) chloride (28)

Pt

P S

S

NN

P

Cl1

2

3

4

5

6

7 8

6'

7'

3'

2'

ef

a

b

cd




Calculated (Found) for C40H43ClN2P2PtS2 (Mol. Wt: 908.4 g/mol): C, 52.89 (52.84); H,

4.77 (4.74); N, 3.08 (3.05); S, 7.06 (7.03). FT-IR (4000-200 cm-1

): 1529 ν(N-CSS), 1239


Chlorido[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophen-

ylphosphine)]platinum(II) (29)

59

Pt

Cl

P S

S

N

F

N OH1

2

3

3'

2'

4

5 6

7

a

b

cd

F

F

6'5'

Amount of reagents used: 0.276 g (1 mmol) sodium 4-(4-hydroxyphenyl)piperazine-1-




H, 3.15 (3.11); N, 3.50 (3.48); S, 8.01 (7.98). FT-IR (4000-200 cm-1

): 1495 ν(N-CSS),


Chlorido[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-chlorophe-


Pt

Cl

P S

S

NCl

ClCl

N OH1

2

34

5 6

7

6'5'3'

2'

a

b

cd





2.97 (2.95); N, 3.30 (3.28); S, 7.55 (7.52). FT-IR (4000-200 cm-1

): 1510 ν(N-CSS), 1225


Chlorido[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolyl-


Pt

Cl

P S

S

NN OH

12

3

3'

2'

4

5 6

7

a

b

cd

60





N, 3.68 (3.65); S, 8.44 (8.42). FT-IR (4000-200 cm-1

): 1512 ν(N-CSS), 1225 ν(C-NC2),


Chlorido[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphos-


Pt

Cl

P S

S

NN OH

12

3

3'

2'

4

5 6

7

a

b

cd

6'5'





N, 3.55 (3.52); S, 8.14 (8.12). FT-IR (4000-200 cm-1

): 1512 ν(N-CSS), 1225 ν(C-NC2),


4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(1,4-bis(diphenylphosphin-

o)butane)platinum(II) chloride (33)

Pt

P S

S

NN

P

ClOH1

2

3

2'

3'

4

5 6

7

6'5'

a

b

cd

ef




61


4.54 (4.52); N, 3.08 (3.05); S, 7.04 (7.02). FT-IR (4000-200 cm-1

): 1512 ν(N-CSS), 1225


Chlorido(morpholine-4-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platinum

(II) (34)

Pt

Cl

P S

S

NO

12

3

a

b

cd

Amount of reagents used: 0.185 g (1 mmol) sodium morpholine-4-carbodithioate, 0.266

g (1 mmol) platinum(II) chloride and 0.276 g (1 mmol) diphenyl-p-tolylphosphine.

Yield: 0.51 g, 77%. m.p. 241 C. Elemental analysis, % Calculated (Found) for

C24H25ClNOPPtS2 (Mol. Wt: 669.1 g/mol): C, 43.08 (43.03); H, 3.77 (3.74); N, 2.09

(2.06); S, 9.58 (9.55). FT-IR (4000-200 cm-1

): 1527 ν(N-CSS), 1244 ν(C-NC2), 1024

ν(C-S), 361 ν(Pt-S), 304 ν(Pt-Cl), 227 ν(Pt-P).

Chlorido(morpholine-4-carbodithioato-κ2S,Sʹ)(tris-p-methoxyphenylphosphine)]-

platinum(II) (35)

Pt

Cl

P S

S

N

H3CO

O

12

3

a

b

cd

OCH3

H3CO

Amount of reagents used: 0.185 g (1 mmol) sodium morpholine-4-carbodithioate, 0.266

g (1 mmol) platinum(II) chloride and 0.352 g (1 mmol) tris-p-methoxyphenylphosphine.

Yield: 0.53 g, 71%. m.p. 247 C. Elemental analysis, % Calculated (Found) for

C26H29ClNO4PPtS2 (Mol. Wt: 745.2 g/mol): C, 41.91 (41.88); H, 3.92 (3.90); N, 1.88

(1.84); S, 8.61 (8.58). FT-IR (4000-200 cm-1

): 1498 ν(N-CSS), 1249 ν(C-NC2), 1019


62

Chlorido(piperidine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platinum-

(II) (36)

Amount of reagents used: 0.183 g (1 mmol) sodium piperidine-1-carbodithioate, 0.266

g (1 mmol) platinum(II) chloride and 0.276 g (1 mmol) diphenyl-p-tolylphosphine.

Yield: 0.5 g, 75%. m.p. 243-245 C. Elemental analysis, % Calculated (Found) for

C25H27ClNPPtS2 (Mol. Wt: 667.1 g/mol): C, 45.01 (44.96); H, 4.08 (4.05); N, 2.10

(2.06); S, 9.61 (9.58). FT-IR (4000-200 cm-1

): 1540 ν(N-CSS), 1246 ν(C-NC2), 999 ν(C-

S), 375 ν(Pt-S), 297 ν(Pt-Cl), 235 ν(Pt-P).

Chlorido(piperidine-1-carbodithioato-κ2S,Sʹ)(tris-p-methoxyphenylphosphine)]plat-

inum(II) (37)

Pt

Cl

P S

S

N

H3CO

1 2

2'

3

3'

4

OCH3

H3CO

a

b

cd

Amount of reagents used0.183 g (1 mmol) sodium piperidine-1-carbodithioate, 0.266 g

(1 mmol) platinum(II) chloride and 0.352 g (1 mmol) tris-p-methoxyphenylphosphine.


C27H31ClNO3PPtS2 (Mol. Wt: 743.2 g/mol): C, 43.64 (43.61); H, 4.20 (4.16); N, 1.88

(1.84); S, 8.63 (8.60). FT-IR (4000-200 cm-1

): 1498 ν(N-CSS), 1249 ν(C-NC2), 1019


Chlorido(4,4’-trimethylenedipiperidine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophen-

ylphosphine)platinum(II) (38)

Pt

Cl

P S

S

N1 2

2'

3

3'

4

a

b

cd

63

Pt

Cl

P S

S

NF

FF

(CH2)3

Pt

Cl

PS

S

NF

FF

12

3

2'

3'

1'2' '

2' ' '

3' '

3' ' '

a

b

cd

Amount of reagents used: 0.406 g (1 mmol) sodium- , ’-trimethylenedipiperidine-1-


flourophenylphosphine. Yield: 1.02 g, 70%. m.p. 233-235 C. Elemental analysis, %

Calculated (Found) for C51H48Cl2F6N2P2Pt2S4 (Mol. Wt: 1454.2 g/mol): C, 42.12 (42.10);

H, 3.33 (3.29); N, 1.93 (1.90); S, 8.82 (8.80). FT-IR (4000-200 cm-1

): 1495 ν(N-CSS),


Chlorido(4,4’-trimethylenedipiperidine-1-carbodithioato-κ2S,Sʹ)(tris-p-chlorophen-

ylphosphine)platinum(II) (39)

Pt

Cl

P S

S

NCl

ClCl

(CH2)3

Pt

Cl

PS

S

NCl

ClCl

12

3

2'

3'

1'2' '

2' ' '

3' '

3' ' '

a

b

cd



chlorophenylphosphine. Yield: 1.1 g, 71%. m.p. 224-226 C. Elemental analysis, %

Calculated (Found) for C51H48Cl8N2P2Pt2S4 (Mol. Wt: 1552.9 g/mol): C, 39.44 (39.40);

H, 3.12 (3.09); N, 1.80 (1.78); S, 8.26 (8.23). FT-IR (4000-200 cm-1

): 1479 ν(N-CSS),


Chlorido(4,4ʹ-trimethylenedipiperidine-1-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylph-

osphine)platinum(II) (40)

Pt

Cl

P S

S

N(CH2)3

Pt

Cl

PS

S

N

12

3

2'

3'

1'2' '

2' ' '

3' '

3' ' '

a

b

cd

64




(Found) for C51H56Cl2N4P2Pt2S4 (Mol. Wt: 1376.3 g/mol): C, 44.51 (44.48); H, 4.10

(4.05); N, 4.07 (4.05); S, 9.32 (9.31). FT-IR (4000-200 cm-1

): 1535 ν(N-CSS), 1252 ν(C-

NC2), 1020 ν(C-S), 349 ν(Pt-S), 297 ν(Pt-Cl), 235 ν(Pt-P).

Chlorido(4,4ʹ-trimethylenedipiperidine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphos-


Pt

Cl

P S

S

N(CH2)3

Pt

Cl

PS

S

N

12

3

2'

3'

1'2' '

2' ' '

3' '

3' ' '

a

b

cd




(Found) for C55H64Cl2N4P2Pt2S4 (Mol. Wt: 1432.4 g/mol): C, 46.12 (46.09); H, 4.50

(4.49); N, 3.91 (3.89); S, 8.95 (8.93). FT-IR (4000-200 cm-1

): 1531 ν(N-CSS), 1254 ν(C-

NC2), 1018 ν(C-S), 361 ν(Pt-S), 301 ν(Pt-Cl), 217 ν(Pt-P).

(4,4ʹ-trimethylenedipiperidine-1-carbodithioato-κ2S,Sʹ)(1,4-bis(diphenylphosphino)-


Pt

P S

S

N

P

Cl2Pt

PS

S

N

P

12

3

2'

3'

1'2' '

2' ' '3' '

3' ' '

a

b

c

d

ef

4

4'5 5'

6




65

Calculated (Found) for C71H80Cl2N2P4Pt2S4 (Mol. Wt: 1674.6 g/mol): C, 50.92 (50.90);

H, 4.82 (4.81); N, 1.67 (1.65); S, 7.66 (7.63). FT-IR (4000-200 cm-1

): 1537 ν(N-CSS),

1258 ν(C-NC2), 1026, 997 ν(C-S), 347 ν(Pt-S), 239 ν(Pt-P).

Thiocyanato[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flouro-

phenylphosphine)]platinum(II) (29a)

Pt

NCS

P S

S

N

F

N OH1

2

3

3'

2'

4

5 6

7

a

b

cd

F

F

6'5'

Amount of reagents used: 0.8 g (1 mmol) chlorido[4-(4-hydroxyphenyl)piperazine-1-

carbodithioato-κ2S,Sʹ)(tris-p-flourophenylphosphine)]platinum(II) and 0.081 g (1 mmol)

sodium thiocyanate. Yield: 0.65 g, 79%. m.p. 241-244 C. Elemental analysis, %

Calculated (Found) for C30H25F3N3OPPtS3 (Mol. Wt: 822.8 g/mol): C, 43.79 (43.74); H,

3.06 (3.03); N, 5.11 (5.08); S, 11.69 (11.67). FT-IR (4000-200 cm-1

): 2055br, 2113sh

ν(NCS), 1495 ν(N-CSS), 1227 ν(C-NC2), 1013 ν(C-S), 368 ν(Pt-S), 230 ν(Pt-P).

Bromido[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophe-

nylphosphine)]platinum(II) (29b)

Pt

Br

P S

S

N

F

N OH1

2

3

3'

2'

4

5 6

7

a

b

cd

F

F

6'5'



ammonium bromide. Yield: 0.7 g, 84%. m.p. 228-230 C. Elemental analysis, %

Calculated (Found) for C29H25BrF3N2OPPtS2 (Mol. Wt: 844.6 g/mol): FT-IR (4000-200

cm-1

): 1495 ν(N-CSS), 1226 ν(C-NC2), 1013 ν(C-S), 381 ν(Pt-S), 209 ν(Pt-Br), 223 ν(Pt-

P).

66

Iodo[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-flourophenyl-

phosphine)]platinum(II) (29c)



potassium iodide. Yield: 0.74 g, 82%. m.p. 234-234 C. Elemental analysis, %

Calculated (Found) for C29H25F3IN2OPPtS2 (Mol. Wt: 891.6 g/mol): FT-IR (4000-200

cm-1

): 1494 ν(N-CSS), 1227 ν(C-NC2), 1012 ν(C-S), 387 ν(Pt-S), 181 ν(Pt-I), 224 ν(Pt-

P).

Thiocyanato[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tri-p-tolyl-

phosphine)]platinum(II) (32a)

Pt

NCS

P S

S

NN OH

12

3

3'

2'

4

5 6

7

a

b

cd

6'5'


carbodithioato-κ2S,Sʹ)(tris-p-tolylphosphine)]platinum(II) and 0.081 g (1 mmol) sodium

thiocyanate. Yield: 0.63 g, 78%. m.p. 242-244 C. Elemental analysis, % Calculated

(Found) for C33H34N3OPPtS3 (Mol. Wt: 810.9 g/mol): C, 48.88 (48.86); H, 4.23 (4.19);

N, 5.18 (5.16); S, 11.86 (11.81). FT-IR (4000-200 cm-1

): 2050br, 2113sh ν(NCS), 1512

ν(N-CSS), 1226 ν(C-NC2), 1017 ν(C-S), 381 ν(Pt-S), 226 ν(Pt-P).

Bromido[4-(4-hydroxyphenyl)piperazine-1-carbodithioato-κ2S,Sʹ)(tris-p-tolylphos-

phine)]platinum(II) (32b)

Pt

I

P S

S

N

F

N OH1

2

3

3'

2'

4

5 6

7

a

b

cd

F

F

6'5'

67

Pt

Br

P S

S

NN OH

12

3

3'

2'

4

5 6

7

a

b

cd

6'5'


carbodithioato-κ2S,Sʹ)(tris-p-tolylphosphine)]platinum(II) and 0.098 g (1 mmol) ammon-

ium bromide. Yield: 0.67 g, 81%. m.p.236-238 C. Elemental analysis, % Calculated

(Found) for C32H34BrN2OPPtS2 (Mol. Wt: 832.7 g/mol): C, 46.16 (46.12); H, 4.12 (4.07);

N, 3.36 (3.33); S, 7.70 (7.67). FT-IR (4000-200 cm-1

): 1512 ν(N-CSS), 1226 ν(C-NC2),

1018 ν(C-S), 384 ν(Pt-S), 204 ν(Pt-Br), 227 ν(Pt-P).

Thiocyanato(morpholine-4-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylphosphine)]plati-

num(II) (34a)

Amount of reagents used: 0.669 g (1 mmol) chlorido(morpholine-4-carbodithioato-

κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platinum(II) and 0.081 g (1 mmol) sodium thiocyan-

ate. Yield: 0.595 g, 86%. m.p. 235-238 C. Elemental analysis, % Calculated (Found) for

C25H25N2OPPtS3 (Mol. Wt: 691.7 g/mol): C, 43.41 (43.38); H, 3.64 (3.60); N, 4.05

(4.02); S, 13.91 (13.89). FT-IR (4000-200 cm-1

): 2052, 2108 ν(NCS), 1524 ν(NCSS),

1243 ν(C-NC2), 1024 ν(C-S), 380 ν(Pt-S), 238 ν(Pt-P).

Bromido(morpholine-4-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platin-

um(II) (34b)

Pt

Br

P S

S

NO

12

3

a

b

cd

Pt

NCS

P S

S

NO

12

3

a

b

cd

68


κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platinum(II) and 0.098 g (1 mmol) ammonium

bromide. Yield: 0.567 g, 80%. m.p. 242-244 C. Elemental analysis, % Calculated

(Found) for C24H25BrNOPPtS2(Mol. Wt: 713.6 g/mol): FT-IR (4000-200 cm-1

): 1527

ν(N-CSS), 1243 ν(C-NC2), 1024 ν(C-S), 384 ν(Pt-S), 210 ν(Pt-Br), 223 ν(Pt-P).

Iodo(morpholine-4-carbodithioato-κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platinum(II)

(34c)

Pt

I

P S

S

NO

12

3

a

b

cd


κ2S,Sʹ)(diphenyl-p-tolylphosphine)]platinum(II) and 0.166 g (1 mmol) potassium iodide.


C24H25INOPPtS2 (Mol. Wt: 760.6 g/mol): FT-IR (4000-200 cm-1

): 1525 ν(N-CSS), 1242

ν(C-NC2), 1024 ν(C-S), 384 ν(Pt-S), 179 ν(Pt-I), 225 ν(Pt-P).

2.5 Computational studies

All the quantum chemical calculations were carried out using GAUSSIAN 09 set of

programs and the results were analyzed via GAUSSVIEW software [4, 5]. Geometries of

the complexes were optimized at DFT/B3LYP level with LANL2DZ basis set. Mullikan

and natural population were obtained at same level of the theory [6, 7]. Molecular

properties like dipole moment, frontier molecular orbitals (HOMO & LUMO), HOMO–

LUMO band gap (ΔE), ionization potential (IP), electron affinity (EA), global hardness

(η), chemical potential (μ) and global electrophilicity (ω) were assessed after DFT

optimization. Global hardness and chemical potential were calculated from the following

equation:

= 1/2(LUMO HOMO) (1)

µ = 1/2(LUMO + HOMO) (2)

69

2.6 DNA-binding using UV-visible spectroscopy

The CT-DNA (20 mg) was dissolved in double deionized water to prepare stock solution

and then its concentration was determined spectrophotometrically at 260 nm using

epsilon value of 6600 M-1

cm-1

[8]. The value of A260/A280 ~1.9 confirmed protein free

nature of the DNA. The UV absorption titrations were performed by keeping

concentration of the prepared complexes fixed and varying the DNA concentration.

During titration, same DNA concentrations were added to both reference and sample

cuvettes to eliminate the absorbance of DNA itself. After ~5 min of the solutions mixing,

absorption spectra were recorded at room temperature in a range of 200-800 nm. The

titration procedures were repeated until there was no change in the spectra for four

titrations at least; indicating binding saturation had been achieved. Benesi–Hildebrand

equation [9] was used to determine the association/binding constants.

=

Where K is the association/binding constant, Aₒ and A are the absorbances of the

complex and its adduct with DNA respectively, and ɛG and ɛH–G are the absorption

coefficients of the compound and the compound-DNA adduct, respectively. The Gibb’s

free energy (ΔG) was determined from the following equation:

ΔG = RT ln K

Where R is the general gas constant (8.314 J K-1

mol-1

) and T is the temperature (298 K).

2.7 DNA-binding using viscosity measurements

Viscosity measurements were carried out using a glass viscometer at room temperature.

A 100 µM solution of the DNA and a series of solutions, containing this constant DNA

concentration with varying concentrations of the complexes (50, 100, 150, 200 and 250

µM), were prepared in DMSO. A digital stopwatch was used to measure the flow time

and an average flow time of three readings was calculated for each solution. First relative

viscosity “ηo” of DNA to solvent was calculated and then relative viscosities (η) of series

of the solutions containing constant DNA and varying concentrations of the complexes

70

were measured. The values of relative specific viscosities (η/(ηo)) were plotted against

[Complex]/[DNA] ratio [10].

2.8 DNA-binding using cyclic voltammetry

Cyclic voltammetric measurements were performed for 2 on Biologic SP-300 cyclic

voltammeter running with EC-lab Express V 5.40 software Japan having three electrodes

system i.e. Pt-disc as working electrode, Pt-wire as counter electrode and Ag/AgCl as

reference electrode. Analytical grade TBAP was used as supporting electrolyte. Before

every reading, working electrode was polished with alumina powder and rinsed with

distilled water [11]. Nitrogen gas (99.9%) was purged through the working solution to

avoid oxygen interference. Drug-DNA binding constant was determined with equation:

log(1/[DNA]) = logK + log(I/Io −I)

Where K is the binding constant, Io and I are the peak currents of free drug and DNA-

bound drug respectively. Following form of Randles–Sevcik equation was used for the

calculation of diffusion coefficients [11]:

Ipa = 2.99 × 105n (αn)

1/2 ACo

Do 1/2

V1/2

Where Ipa is the anodic peak current, Co is the reductant’s concentration in mol cm−

, A is

the geometric area of the electrode in cm2, n is the number of electrons involved in the

process and Do is the diffusion coefficient in cm2

s−

.

2.9 Cytotoxic potential against HepG2 cancer cell line

Human hepatocellular carcinoma (HepG2) cells were maintained in DMEM containing

FBS (10%), Na-pyruvate (1 mM), L-glutamine (2 mM), penicillin (100 µg/ml) and

streptomycin (100 µg/ml) under a humidified atmosphere at 5% CO2 and 37 °C.

Cytotoxicity of the representative complexes (3-5, 7-14, 17-19, 25, 27, 28, 38-42) was

measured by MTT assay [12]. In short, HepG2 cultures (>90% viability; 1.5×105

cells/ml) were exposed to six different concentrations (1, 3, 10, 30, 100 and 300 µM) of

the complexes, doxorubicin and cisplatin for 24 hours. As experimental controls,

unexposed samples, DMSO (solvent) treated and non-cellular background samples i.e.,

71

‘media only’ and ‘compounds only’ were included in each experiment. Then cultures

were incubated with MTT solution (0.5 mg/ml) for 3 hours at 37 °C to produce formazan

crystals which were dissolved in acidified 10% SDS and absorbance was measured at 565

nm by using a microplate reader (AMP PLATOS R-496). Relative percent viability of the

treated samples was calculated using the following formula:

Relative percent viability = [Abs (565)test Abs (565)compound control/Abs (565)unexposed control

Abs (565)media] × 100,

Where Abs (565)test and Abs (565)unexposed control represent the optical density at 565 nm for

the treated samples and untreated control samples respectively. Abs (565)compound control

and Abs (565)media represent background optical density and was measured in compound

only and media only samples respectively. IC50 values for the compounds representing

the concentrations at which 50% of cell growth is inhibited, were calculated.

2.10 Cytotoxic potential against five cancer cell lines

Sulforhodamine B (SRB) cellular protein-staining method [13] was used to evaluate the

cytotoxic potential of the three representative complexes 1, 2 and 6 towards proliferation

of five different human cancer cell lines. Cancer cells (1 104 cells in 190 µl of the

complete media) were plated in 96-well plates containing the tested compounds dissolved

in 100% DMSO and incubated (37 , 5% CO2 in humidified air) for 72 hours. The

incubation was stopped at the end with tri-chloroacetic acid. The cells were washed, air

dried and stained with SRB solution, and after that optical densities (ODs) were measured

at 515 nm using a microplate reader. A zero-day control was performed in each case by

adding an equivalent number of cells to several wells, incubating at 37 for 30 min and

processing as described above. The percentage cell survival was calculated using the

following formula:

2.11 Anticancer activity against LU and MCF-7 cancer cell lines

Cytotoxicity of the complexes was also investigated against LU (human lung carcinoma)

and MCF-7 (human breast adenocarcinoma) by the same MTT procedure as described

72

above, except exposing these cell lines herein, to six different concentrations (1, 3, 10,

30, 100, and 300 µM) of the complexes and cisplatin for 48 hours.

2.12 Cleavage of plasmid DNA

Plasmid DNA (pUC 19) cleavage activity of the complexes was monitored by using

agarose gel electrophoresis [14]. In a typical experiment, supercoiled pUC 19 DNA (12.5

μg/ml, μl) in Tris-HCl (100 mM, pH 7.4) was treated with different complex

concentrations, followed by dilution with the mentioned buffer to a total volume of μl.

The samples were then incubated at 37 for 6 hours and loaded on a 0.7% agarose gel

containing . μg/ml ethidium bromide. Electrophoresis was carried out at 40 V for 30

min in TAE buffer and run in duplicate. DNA-bands were visualized in UV light and then

photographed followed by estimation of their intensity using a gel documentation system.

73

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1. Armarego, W. L.; Chai, C. L. L., Purification of laboratory chemicals,

Butterworth-Heinemann 2013.

2. Armarego W.L.; C.L.L. Chai, Purification of Laboratory Chemicals, 7th Edition,

Butterworth-Heinemann, Elsevier Inc., Burlington 2009.

3. Sheldrick, G. M., A short history of SHELX, Acta Crystallogr. Sect. A, 2008, 64,

112-122.

4. Frisch, M.; Trucks, G.; Schlegel, H. B.; Scuseria, G.; Robb, M.; Cheeseman, J.;

Scalmani, G.; Barone, V.; Mennucci, B.; Petersson, G., Gaussian 09, revision a.

02, gaussian. Inc., Wallingford, CT 2009, 200.

5. Frisch, A.; Nielsen, A.; Holder, A., GAUSSIANVIEW Users Manual, Gaussian

Inc., Pittsburgh PA, 2000.

6. Abdel-Ghani, N.; Mansour, A., Molecular structures of antitumor active Pd (II)

and Pt (II) complexes of N, N-donor benzimidazole methyl ester, J. Coord. Chem.

2012, 65, 763-779.

7. Zhang, G.; Musgrave, C. B., Comparison of DFT methods for molecular orbital

eigenvalue calculations, J. Phys. Chem. A 2007, 111, 1554-1561.

8. Reichmann, M.; Rice, S.; Thomas, C.; Doty, P., A further examination of the

molecular weight and size of desoxypentose nucleic acid, J. Am. Chem. Soc.

1954, 76, 3047-3053.

9. Shujha, S.; Shah, A.; Muhammad, N.; Ali, S.; Qureshi, R.; Khalid, N.; Meetsma,

A., Diorganotin (IV) derivatives of ONO tridentate Schiff base: synthesis, crystal

structure, in vitro antimicrobial, anti-leishmanial and DNA binding studies, Eur.

J. Med. Chem. 2010, 45, 2902-2911.

10. Shi, S.; Liu, J.; Li, J.; Zheng, K.-C.; Huang, X.-M.; Tan, C.-P.; Chen, L.-M.; Ji,

L.-N., Synthesis, characterization and DNA-binding of novel chiral complexes Δ-

and Λ-[Ru(bpy)2L]2+

(L = o-mopip and p-mopip), J. Inorg. Biochem. 2006, 100,

385-395.

11. Holze, R., CMA Brett, AMO Brett: Electrochemistry-Principles, methods and

applications, Oxford University Press, Oxford, ISBN 0‐19‐855388‐9, 1993, 427.

74

12. Mosmann, T., Rapid colorimetric assay for cellular growth and survival:

application to proliferation and cytotoxicity assays, J. Immunol. Methods 1983,

65, 55-63.

13. You, M.; Wickramaratne, D. M.; Silva, G. L.; Chai, H.; Chagwedera, T. E.;

Farnsworth, N. R.; Cordell, G. A.; Kinghorn, A. D.; Pezzuto, J. M., (-)-

Roemerine, an aporphine alkaloid from Annona senegalensis that reverses the

multidrug-resistance phenotype with cultured cells, J. Nat. Prod. 1995, 58, 598-

604.

14. Raman, N.; Thalamuthu, S.; Dhaveethuraja, J.; Neelakandan, M.; Banerjee, S.,

DNA cleavage and antimicrobial activity studies on transition metal (II)

complexes of 4-aminoantipyrine derivative, J. Chil. Chem. Soc. 2008, 53, 1439-

1443.

Chapter 3

RESULTS AND DISCUSSION

75

3.1 FT-IR

FT-IR data of the ligands (1-10) and their Pt(II) complexes (1-42) are given in the

experimental section. Metal dithiocarbamates give distinctive bands for C-N (1450-1550

cm-1

), C-S (950-1050 cm-1

) and M-S (300-400 cm-1

) bonds [1]. Generally, stretching

frequencies for C-N and C=N bonds appear in the range 1200-1350 cm-1

and 1640-1690

cm-1

, respectively [2,3]. In dithiocarbamate, the C N stretch appears at 1430-1550 cm-1

[1, 4], which is concomitant with its partial double bond nature. In ligands, the

appearance of C N stretch at 1433-1474 cm-1

indicates considerable contribution of the

resonance form I (Fig. 3.1). This band shifts to higher frequency (1479-1540 cm-1

) upon

complexation, thus signifying an increase in the bond order owing to high contribution

from canonical form IV (Fig. 3.1).

N CS

SN C

S

S

N C

S

S

N C

S

S

I II III IV

Figure 3.1: Resonance forms of the dithiocarbamic-NCSS moiety.

In ligands, the appearance of two peaks (1007-1047 cm-1

and 985-1003 cm-1

) for υ(C-S)

expressing unsymmetrical nature of the –NCSS group imposed by the higher contribution

from the resonance forms II and III (Fig. 3.1). However, for complexes manifestation of

a single peak at 997-1030 cm-1

for the aforesaid bond shows bidentate coordination of –

NCSS group via its two sulfur atoms. New bands at 345-390 cm-1

and 211-240 cm-1

correspond to Pt-S and Pt-P bonds show the attachment of dithiocarbamate and

organophosphine to the platinum center [1, 5]. The Pt-Cl stretch (273-316 cm-1

) was

noted in the monofunctional complexes only [6], and hence indicates the replacement of a

single chloride by the anionic dithiocarbamate ligand.

3.2 1H NMR

1H NMR spectra of the ligands and their complexes were recorded in DMSO-d6 and

CDCl3 respectively. Assignments of 1H-resonances were made by peak multiplicity,

intensity pattern and comparison of integration values with the expected composition.

76

Presence of proton resonances for both dithiocarbamate ligands and organophosphines

signify formation of the complexes. 1H NMR data of the ligands (1-10) and their Pt(II)

complexes are presented in Tables 3.1-3.5. In the ligands 1-4 and 6-9, methylene protons

of piperazine ring at position 2, 2´ were appeared as triplet in the range 4.14-4.45 ppm (J

= 4-5.4 Hz). In other two ligands (5 and 10), they appeared as multiplet at 3.61-3.66 ppm

and doublet at 5.78 ppm (J = 12 Hz) respectively. However, in the complexes, the

piperazine protons (position 2, 2´-Hs) appeared in diverse manner, i.e. as doublets in 39-

42 (4.18-4.24 ppm J = 12 Hz), as triplets in 3-5, 16-19, 28, 31-33 (3.46-4.00 ppm, J = 4-

5.7 Hz), as multiplets in 1-2, 6-7, 11-15, 20-27, 30, 34-38 (2.59-4.48 ppm) and as broad

signals in 8-10 (3.86-3.95 ppm). The upfield shift of the signals in complexes indicates

complexation of the dithiocarbamate ligands and back-donation of electron density from

Pt(II) in the presence of electron donating organophosphine ligand [7]. Chemical shifts of

all other protons of the ligands (dithiocarbamate and organophosphines) were observed

identical to the free ligands. 1H NMR spectra of a representative ligand (L1) and its

corresponding monofunctional Pt(II) complex (2) are shown in the Fig. 3.2 & 3.3

respectively. In comparison, appearance of the new peaks of aromatic region at 7.42-7.62

ppm in the complex spectrum indicates the presence of organophosphine in the structure.

3.3 13

C NMR

13C NMR spectra of the ligands and their complexes were recorded in DMSO-d6 and

CDCl3, respectively. 13

C NMR data of the free ligands (1-10) and their Pt(II) complexes

are collected in Tables 3.6-3.10. In complexes, a significant 13

C-shift was observed

compared to corresponding free ligands, in the area closer to the metal coordination site.

In ligands (1-10), the chemical shift for CS2 carbon appeared in range 212.7-214.9 ppm

while in complexes (1-42) at 197.6-211.0 ppm. The upfield shift in complexes can be

attributed to an increase in the electron density on 1,1-dithioate carbon due to

mobilization of the nitrogen lone pair of NCSS toward CSS moiety on complexation; an

observation consistent with the IR data [8]. Back-donation of the electron density from

Pt(II) to dithiocarbamate in the presence of electron donating organophosphine may also

has contributed in the upfield shift. The carbon at position 2, 2´ of piperazine ring in most

of the complexes is shifted upfield which can be attributed to the back donation and as

well as to stronger binding of dithiocarbamate to the Pt(II) than the chloride [4].

77

Nevertheless, aromatic carbons of organophosphine appeared as doublet {Fig. 3.5

(inset)} due to phosphorous platinum coupling [9]. All other signals in 13

C NMR spectra

of the ligands and their complexes were observed at almost identical positions. 13

C NMR

spectra of a representative ligand (L1) and its corresponding monofunctional Pt(II)

complex (2) are shown in the Figs. 3.4 & 3.5, indicating a significant upfield shift of CSS

peak upon complexation. Moreover, appearance of new peaks in the aromatic region

(126.6-138.0 ppm) indicates the presence of organophosphine in the structure.

3.4 31

P NMR

31P NMR spectra of the Pt(II) complexes (1-42) were recorded in CDCl3 and are

presented in the Tables 3.6-3.10. Chemical shifts of the complexes were observed in the

range 9.49-15.35 ppm indicating downfield shifts compared to the free

organophosphines, thus confirming the coordination with platinum centre [10]. 31

P NMR

spectrum of the representative monofunctional Pt(II) complex (2) is shown in the Fig.

3.6.

Figure 3.2: The 1H NMR spectrum of the representative ligand (L1).

78

Figure 3.3: The 1H NMR spectrum of the representative monofunctional Pt(II) complex

(2).

Figure 3.4: The 13

C NMR spectrum of the representative ligand (L1).

79

Figure 3.5: The 13

C NMR spectrum of the representative monofunctional

Pt(II)complex(2).

Figure 3.6: The 31

P NMR spectrum of the representative monofunctional Pt(II) complex

(2).

80

Table 3.1: 1H NMR data of the ligands (L1 and L2) and their Pt(II) complexes (1-10).

“J” values (Hz) are given in brackets, s: singlet, d: doublet, t: triplet, m: multiplet, br: broad

Where X = F (1, 6), Cl (2, 7), CH3 (3, 4, 8, 9)

Comp

. No.

Chemical shift δ (ppm)

Methylene

-CH2 (2, 2′) -CH2( 3, 3′) -CH (5, 6) -OCH3 (8) -CH (b-c) -CH3 -CH2-CH2-CH2-CH2-

L-1 4.45, t (5.1) 2.93, t (5.1) 6.80-6.92, m 3.67, s - - -

1 3.81-4.01, m 3.09-3.19, m 6.81-6.99, m 3.78, s 7.07-7.72, m - -

2 3.81-4.02, m 3.07-3.14, m 6.85-6.92, m 3.79, s 7.42-7.64, m - -

3 3.88, t (4.5) 3.14, t (4.5,) 6.83-6.92, m 3.78, s 7.01-7.71, m 2.40, s -

4 3.89, t (4.8) 3.14, t (4.8,) 6.81-6.96, m 3.77, s 7.07-7.59, m 2.39, s -

5 3.77, t (4) 3.01, t (4) 6.70-6.80, m 3.66, s 7.40-7.47, m - 1.92-2.11, m

-CH (6) -CH (7) -CH (8)

L-2 4.39, t (5.1) 3.62, s(br) 7.02, d (3.6) 6.62, dd 7.84, d - - -

6 3.90-3.96, m 3.75-3.79, m 7.86, s(br) 6.65, dd 7.62, d 7.10-7.72, m - -

7 3.91-3.96, m 3.76-3.79, m 7.14, d (3.7) 6.54, dd 7.50, d 7.35-7.63, m - -

8 3.86, s(br) 3.65-3.79, m 7.03-7.62, m 6.44, dd 7.62, d 7.03-7.62, m 2.32, s -

9 3.95, s(br) 3.75-3.87, m 7.13, d (3.3) 6.52, dd 7.22, d 7.25-7.59, m 2.40, s -

10 3.94, s(br) 3.85, t (3) 7.15, d (3.3) 6.51, dd 7.50, s(br) 7.53-7.58, m

2.01-2.19, m

81



Where X = F (11, 16), Cl (12), CH3 (13, 14, 17, 18)

Comp. No.


Methylene

-CH2 (2, 2′) -CH

2( 3, 3′) -CH (4) -CH (6-8, b-c) -CH

3 -CH

2-CH

2-CH

2-CH

2-

L-3 4.32, t (4.8) 2.22, t (4.8) 4.29, s 7.16-7.44, m - -

11 3.63-3.82, m 2.41-2.50, m 4.26, s 7.08-7.67, m - -

12 3.66-3.86, m 2.44-2.53, m 4.29, s 7.19-7.62, m - -

13 3.62-3.84, m 2.80-3.32, m 4.32, s 7.01-7.75, m 2.30, s -

14 3.57-3.74, m 2.40-2.57, m 4.23, s 6.86-7.53, m 2.29, s -

15 3.21-3.73, m 2.38-2.70, m 4.33, s 7.15-7.83, m(br) - 2.00-2.19, m

-CH (5) -CH (6) -CH (b-c) -CH3

L-4 4.44, t (5.4) 3.49, t (5.4) 6.97, d (9.6) 8.05, d (9.6) - - -

16 3.96, t (5.2) 3.78, t (5.2) 6.77, d (9.2) 8.08, d (9.2) 6.86-7.62, m - -

17 4.00, t (5.7) 3.65, t (5.7) 6.79, d (9.3) 8.10, d (9.3) 6.90-7.70, m 2.34, s -

18 3.94, t (4) 3.60, t (4) 6.87, d (9.3) 8.03, d (9.3) 6.99-7.19, m 2.31, s -

19 3.89, t (5.2) 3.54, t (5.6) 6.81, d (9.2) 8.01, d (9.6) 7.31-7.65, m - 1.93-1.99, m

82


C.No Chemical shift δ (ppm)

Methylene

-CH2 (2, 2′) -CH

2( 3, 3′) -CH

2 (4) -CH

2 (5) -OH -CH (b-c) -CH

3 -CH

2-CH

2-CH

2-CH

2-

L-5 3.61-3.66, m 2.45-2.48, m 3.22-3.23, m 4.37, t (4) 1.79, s - - -

20 3.60-3.82, m 2.54-2.61, m 2.98, t (4) 3.92, t (4) 2.11, s 7.01-7.62, m - -

21 3.62-3.69, m 2.55-2.66, m 3.38, t (4) 3.84, t (4) 2.11, s 7.11-7.58, m - -

22 3.59-3.84, m 2,51-2.58, m 3.12-3.16. m 3.92, s (br) 2.11, s 6.99-7.17, m 2.31, s -

23 3.64-3.81, m 2.58-3.04, m 3.00, t (4) 3.87, t (4) 2.11, s 6.99-7.17, m 2.31, s -

24 2.59-2.75, m(br) 2.50-2.58, m 1.96, t(8) 3.66, (4) 1.18, s 7.22-7.73, m - 1.64-1.80, m

,

-CH2 (4) -CH (6-8, b-c) -CH

3

L-6 4.30, t (4.8) 2.27, t (4.8) 3.45, s 7.22-7.37, m - -

25 3.63-3.82, m 2.41-2.50, m 3.50, s 6.86-7.66, m - -

26 3.61-3.84, m 2.40-2.57, m 3.51, s 6.89-7.65, m 2.30, s -

27 3.56-3.83, m 2.37-2.52, m 3.52, s 6.93-7.61, m 2.32, s -

28 3.46, t (4) 2.43, t (4) 4.46, s 7.17-7.49, m - 1.93-2.11, m


Where X = F (20, 25), Cl (21), CH3 (22, 23, 26, 27)

83



Methylene

-CH2 (2, 2′) -CH

2( 3, 3′) -CH (5, 6) -OH -CH (b-c) -CH

3 -CH

2-CH

2-CH

2-CH

2-

L-7 4.45, t (5.1) 2.63, t (5.1) 6.62, d (8), 7.58, d (8) 1.8, s - - -

29 3.67-3.85, m 2.95, s(br) 6.77, d (8), 7.59, d (8) 1.94, s(br) 7.01-7.37, m - -

30 3.71-3.90, m 2.94-3.05, m 6.66, d (8), 7.55, d (8) 1.56, s(br) 6.75-7.51, m - -

31 3.70, t (4) 2.90, t (4) 6.61, d (8), 7.51, d (8) 2.73, s 7.01-7.35, m 2.28, s -

32 3.74, t (4) 2.95, t (4) 6.64, d (8), 7.45, d (8) 1.59, s 6.96-7.19, m 2.31, s -

33 3.72, t (4) 2.93, t (4) 6.65, d (8), 7.66, d (8) 1.67, s 7.24-7.47, m - 1.93-1.98, m(br

-CH2 (2, 2′) -CH

2( 3, 3′) -CH (b-c) -OCH

3 -CH

3

L-8 4.26, t (4) 3.64, t (4) - - - -

34 3.60-3.78, m 3.60-3.78, m 6.89-7.57, m - 2.32, s -

35 3.71-3.78, m 3.71-3.78, m 6.59-7.33, m 1.68, s - -

“J” values (Hz) are given in brackets, s: singlet, t: triplet, m: multiplet, br: broad

Where X = F (29), Cl (30), CH3 (31, 32, 34), OCH3 (35)

84



Methylene

-CH2 (2, 2′) -CH

2( 3, 3′), -CH( 4) -CH (b-c) -OCH3 -CH

3 -CH

2-CH

2-CH

2-CH

2-

L-9 4.14, t (4) 1.54-1.65, m - - - -

36 3.63-3.96, m 1.60-2.02, m 7.02-7.67, m - 2.32, s -

37 3.48-3.74, m 1.57-1.87, m 6.60-7.49, m 1.69, s - -

-CH2 (2, 2′) -CH

2( 3, 3′) -CH (4) -CH

2( 5) -CH

2( 6) -CH (b-c) -CH

3

L-10 5.78, m 2.73-2.79, m 1.28-1.46, m 0.90-1.00, m 1.14-1.19, m - - -

38 4.10-4.48, m 2.87-2.96, m 1.69-1.81, m 1.03-1.19, m 1.55, s(br) 6.87-7.65, m - -

39 4.24-4.44, m 2.96-3.10, m 1.77-1.92, m 1.17-1.30, m 1.67, s(br) 7.31-7.62, m - -

40 4.24-4.44, m 2.98-3.26, m 1.69-1.79, m 1.04-1.12, m 1.13-1.22, m(br) 7.02-7.62, m 2.31, s

41 4.18-4.41, m 2.96-3.15, m 1.74-1.80, m 1.04-1.10, m 1.15-1.22, m(br) 6.98-7.54, m 2.30, s -

42 4.18-4.42, m 2.94-3.06, m 1.64-1.82, m 0.99-1.08, m 1.11-1.19, m(br) 7.32-7.67, m 1.93-1.99, m


Where X = F (38), Cl (39), CH3 (36, 40, 41), OCH3 (37)

85

Table 3.6: 13

C NMR and 31

P NMR data of the ligands (L1 and L2) and their Pt(II) complexes (1-10).

C.No


Dithiocarbamate moiety Organophosphine moiety

Methylene

Carbon Phosphorus

C (1) C (2,2′) C ( 3,3′) C (4) C (5, 6) C (7) C (8) C (a) C (b-c) C (d) CH

3 C (e) C (f)

31P

L-1 214.6 50.3 49.3 145.9 118.0, 114.7 153.4 55.6 - - - - - - -

1 201.0 50.5 47.6 144.3 119.5, 114.5 155.0 55.5 166.3 135.5,116.3 163.0 - - - 13.05

2 205.4 50.5 47.0 144.3 119.5, 114.7 155.0 55.6 127.0 135.4,128.9 137.9 - - - 13.29

3 201.3 50.4 47.2 144.0 119.5, 114.7 155.0 55.6 127.7 134.0,129.6 142.7 21.5 - - 14.47

4 201.8 50.4 47.2 142.3 119.5, 114.6 155.0 55.6 129.0 134.2,129.2 142.3 21.5 - - 15.35

5 201.5 49.3 46.2 143.0 118.4, 113.6 153.9 54.5 127.6 131.9,128.2 131.2 - 22.3 28.3 10.42

C (4) C (5) C (6) C (7) C (8)

L-2 214.9 49.2 40.8 159.0 147.4 116.1 111.8 145.2 - - - - - - -

6 207.2 46.8 46.6 158.9 147.2 118.2 111.9 144.3 166.2 136.4,116.0 162.8 - - - 9.59

7 207.0 46.8 46.7 158.9 147.2 118.2 111.9 144.3 126.8 135.4,129.0 138.0 - - - 10.54

8 207.8 46.9 46.6 159.0 147.2 118.1 111.8 144.3 125.4 134.4,129.2 141.4 21.5 - - 11.03

9 208.2 53.8 46.5 159.0 147.2 118.0 111.8 144.3 126.0 134.2,129.1 141.1 21.5 - - 9.69

10 204.2 46.9 46.8 159.1 146.9 118.3 111.9 144.5 128.6 132.9,129.2 132.2 - 23.0 31.6 -

Where X = F (1, 6), Cl (2, 7), CH3 (3, 4, 8, 9)

C

S

SNa1

86

Table 3.7: 13

C NMR and 31


C.No



Methylene

Carbon Phosphorus

C (1) C (2,2′) C (3,3′) C (4) C (5) C (6, 7) C (8) C (a) C (b-c) C (d) CH

3 C (e) C (f)

31P

L-3 214.4 52.1 49.4 75.2 143.1 128.9, 128.1 127.3 - - - - - - -

11 199.0 50.7 48.1 75.4 141.1 128.6, 128.3 127.2 165.9 135.5,116.5 163.3 - - - 13.21

12 204.5 50.7 47.1 75.5 141.3 128.8, 127.7 127.5 127.0 135.4,128.9 137.9 - - - 10.88

13 200.6 50.6 48.0 76.6 142.7 128.8, 127.7 128.4 127.5 134.2,129.5 141.1 21.5 - - 14.51

14 201.8 50.4 47.2 76.3 141.5 128.5, 127.9 127.3 128.8 134.2,129.2 142.1 21.5 - - 13.31

15 201.9 48.4 43.7 75.5 141.9 128.3, 130.1 127.3 128.5 132.9,129.2 132.5

23.3 26.3 10.48

C (4) C (5, 6) C (7)

L-4 214.6 48.6 46.3 154.9 112.4, 126.2 136.8 - - - - - - -

16 207.0 46.3 45.8 153.7 113.6, 126.0 139.8 165.7 136.4,116.0 163.2 - - - 9.49

17 201.7 46.3 41.0 153.7 113.4, 126.0 142.7 127.6 134.2,129.6 139.3 21.5 - - 14.53

18 202.0 53.8 45.9 153.7 113.3, 126.0 140.9 128.8 134.1,129.3 142.3 21.5 - - 13.31

19 203.1 46.3 46.0 153.6 113.5, 126.0 139.1 128.7 132.8,129.3 132.2 - 23.4 26.6 10.33

Where X = F (11, 16), Cl (12), CH3 (13, 14, 17, 18)

C

S

SNa1

87

Table 3.8: 13

C NMR and 31




Methylene

Carbon

Phosphorus

C (1) C (2,2′) C (3,3′) C(4) C (5) C (a) C (b-c) C (d) CH

3 C (e) C (f)

31P

L-5 213.2 54.4 51.2 61.0 59.8 - - - - - - -

20 211.0 53.8 51.0 59.2 58.0 166.1 136.3,116.6 163.6 - - - -

21 205.7 51.8 46.3 59.2 57.9 127.0 135.1,129.6 137.9 - - - 14.14

22 202.3 51.4 47.5 59.3 57.7 127.5 134.1,129.5 142.4 21.5 - - 14.61

23 200.0 51.7 47.2 59.3 58.5 127.7 134.1,128.7 142.6 21.5 - - -

24 200.3 50.8 46.2 58.4 57.4 127.6 131.8,128.2 131.1 - 22.4 25.6 10.35

C (4) C (5) C (6, 7) C (8)

L-6 214.2 53.2 49.4 62.4 138.4 129.4, 128.6 127.4 - - - - - - -

25 199.3 51.5 47.9 62.6 141.8 128.9, 128.1 127.2 166.0 135.8,116.2 163.2 - - - 12.38

26 200.6 51.6 47.6 62.4 141.9 128.9, 128.1 127.6 126.6 134.6,129.3 141.5 21.5 - - 13.70

27 201.2 51.7 47.2 62.4 142.3 129.1, 128.6 127.6 126.7 134.0,129.3 140.8 21.4 - - 13.30

28 203.9 49.9 44.7 62.5 141.9 129.6, 128.4 127.4 127.9 131.9,129.1 132.3 - 23.5 26.7 10.59

Where X = F (20, 25), Cl (21), CH3 (22, 23, 26, 27)

C

S

SNa1

88

Table 3.9: 13

C NMR and 31


C.No



Methylene

Carbon Phosphorus

C (1) C (2,2′) C (3,3′) C (4) C (5, 6) C (7) C (a) C (b-c) C (d) CH

3 C(e) C(f)

31P

L-7 213.9 50.7 49.6 145.2 111.9, 118.2 150.8 - - - - - - -

29 199.6 50.9 47.4 142.9 115.8, 119.7 152.8 165.9 135.6,116.4 163.3 - - - 13.23

30 205.1 50.6 47.0 143.8 116.2, 119.6 151.7 127.0 135.4,128.9 137.9 - - - 10.97

31 200.8 50.9 47.4 142.7 116.8, 119.6 153.7 127.8 134.3,129.5 142.6 21.5 - - 14.51

32 200.3 49.9 46.3 141.3 115.7, 118.6 152.6 123.5 133.0,128.3 141.3 20.5 - - 13.34

33 202.2 50.9 47.3 142.3 116.7, 119.6 153.6 128.6 132.9,129.2 132.1 - 23.3 34.5 10.44

C (2,2′) C

(3,3′) C (a) C (b-c) C (d) OCH

3 CH

3

L-8 213.4 50.4 69.2 - - - - - - - -

34 201.8 47.4 65.8 127.6 134.3, 129.6 142.7 - 21.5 - - 14.40

35 202.4 47.3 65.8 118.9 135.7, 114.2 162.0 55.7 - - - 11.30

Where X = F (29), Cl (30), CH3 (31, 32, 34), OCH3 (35)

C

S

SNa1

89

Table 3.10: 13

C NMR and 31




Methylene

Carbon

Phosphorus

C (1) C (2,2′) C (3,3ʹ) C (4) C (a) C (b-c) C (d) OCH

3 CH

3 C(e) C(f)

31P

L-9 213.7 51.1 27.3 25.2 - - - - - - - -

36 198.9 48.6 25.6 24.1 127.9 134.3, 129.5 142.6 - 21.5 - - 14.72

37 199.3 48.5 25.5 24.0 119.1 135.6, 114.1 161.9 55.6 - - - 11.52

C (2,2ʹ) C (3,3ʹ) C (4) C (5) C (6) C (a) C (b-c) C (d) CH3

L-10 212.7 49.4 32.2 35.5 36.3 23.3 - - - - - - -

38 203.8 47.3 31.5 35.8 41.0 21.1 165.6 136.5, 115.9 163.1 - - - 10.05

39 197.6 47.5 29.3 31.6 31.8 21.0 127.1 135.4, 128.9 137.8 - - - 14.14

40 202.8 48.0 31.7 35.1 39.0 26.1 128.4 134.3, 129.5 140.3 21.5 - - 14.82

41 198.8 47.8 31.7 35.2 37.0 21.4 124.7 134.1, 129.2 142.1 21.5 - - 13.57

42 199.7 47.9 31.6 35.3 35.8 26.5 128.6 132.9, 129.2 132.1 - 23.4 29.3 10.32

Where X = F (38), Cl (39), CH3 (36, 40, 41), OCH3 (37)

C

S

SNa1

90

3.5 Anticancer study

3.5.1 Anticancer activity against HepG2 cell line

The representative complexes (3-5, 7-14, 17-19, 25, 27, 28 and 38-42) were screened for

in vitro cytotoxic activity against HepG2 cell line by MTT [3-(4,5-dimethylthiazol-2-yl)-

2,5-diphenyltetrazolium bromide] assay using the most effective anticancer drugs

doxorubicin and cisplatin, as standards. The IC50 values (concentration of complex/drug

required to inhibit 50% growth of cancerous cells) are collected in Table 3.11 and their

comparison is presented in Figs. 3.7 & 3.8. The results indicated that about all the

screened monofunctional complexes containing replaceable chloride have activities

greater than that of both the standards. On the other hand, bis-organophosphine

complexes (5, 10, 19, 28 and 42), lacking the labile chloride were found less active than

the standards. Thus, based upon these results an essential role of the replaceable chloride

in the anticancer action can be speculated. These complexes are assumed to remain

integrated in outer cellular environment but get dissociated spontaneously within the cells

due to change in Clˉ ion concentration from high to low. The complexes 4 (IC50 7.9 µM)

and 25 (IC50 38.1 µM) exhibit the highest anticancer activities that are 14 and 3 folds

greater than that of the cisplatin, respectively. Single crystal XRD structure of 25 (Fig.

4.3) and optimized structure of 4 (Table 4.8) show that these complexes are sterically

protected by C-H of aromatic rings (triorganophosphine) from both axial sides. Likewise,

complexes 7, 9, 16-18 and 27 demonstrating greater anticancer activities are also axially

protected though from a single side only. These findings suggest that the axial steric

hindrance provided by aromatic C-H play an important role as it ensures the safe access

of these complexes to the target DNA. Bimetallic complexes (38-41) having replaceable

chloride also show greater anticancer activities than the standards; where complex 41 is

the most active one (IC50 of 23.1 µM), of 5 folds greater activity than the cisplatin. In the

view of structure-activity correlation, higher activities of 4 and 41 may be due to methyl

groups at para positions of the aromatic rings of organophosphines which render them

high lipophilicity to get into the cells. Similarly, higher activities of 11 and 25 may be

because of presence of fluoro moieties, which not only render them high lipophilicity to

facilitate cell internalization [11-13], but also have the potential to stabilize complex-

DNA adducts through hydrogen bonding.

91

Table 3.11: IC50 values of the selected Pt(II) complexes against HepG2 cell line.

Complex IC50 (µM) Complex IC50 (µM)

(3) 112.3 (17) 100.8

(4) 7.9 (18) 60.6

(5) 190.1 (19) 201.2

(7) 54.8 (25) 38.1

(8) 151.2 (27) 55.1

(9) 58.9 (28) 198.2

(10) 200.3 (38) 63.7

(11) 52.7 (39) 65.4

(12) 89.7 (40) 41.0

(13) 63.6 (41) 23.1

(14) 94.1 (42) 160.5

Doxorubicin 127.3 - -

Cisplatin 112.6 - -

Figure 3.7: Comparison of IC50 (µM) values of the representative compounds against

HepG2 cell line.

112.3

7.9

190.1

54.8

151.2

58.9

200.3

52.7

89.7

63.6

94.1 100.1

127.25 112.6

0

50

100

150

200

Compounds

IC5

0 (

µM

)

92

Figure 3.8: Comparison of IC50 (µM) values of the representative compounds against

HepG2 cell line.

3.5.2 Anticancer activity against five different cancer cell lines

In order to envisage the key role of aromatic C-H axial hindrance, three sterically

hindered complexes (complex 1 at both while complexes 2 and 6 at one axial side)

confirmed from single crystal XRD structures (Fig. 4.3), were examined for their in vitro

cytotoxic potential against LU human lung carcinoma, MCF-7 human breast

adenocarcinoma, MDA-MB-231 human breast adenocarcinoma, Hepa-IcIc7 mouse liver

hepatoma and PC-3 human prostate adenocarcinoma by sulforhodamine B (SRB) cellular

protein-staining method using Staurosporine as a standard drug. The IC50 values show

that the complexes (1, 2 and 6) were active against all of the five cancer cell lines (Fig.

3.9 & Table 3.12). The complex 1 showed significant activity against four cell lines (LU,

MDA-MB-231, Hepa-IcIc7 and PC-3) due to the steric hindrance imposed by two axially

leaning aromatic C-H of triorganophosphine. This steric hindrance is expected to stabilize

platinum LUMO orbitals, thus protect it from undergoing spontaneous reactions with the

off-target biomolecules in the intracellular environment. However, this hindrance is

innocent in complex-DNA interaction due to setting up of non-covalent connections

between the adduct molecule, followed by spontaneous reaction between complex and

nitrogen bases of the DNA through LUMO orbital of the former. The relatively low

activity of the complexes 2 and 6 is due to a single sided axial hindrance. The seminal

role of ligand substituents in the anticancer action can also be established. The high

60.6

201.2

38.1 55.1

198.2

63.7 65.4

41

23.1

160.5

127.25 112.6

0

50

100

150

200

Compounds

IC5

0 (

µM

)

93

activity of 1 and 6 may be due to the presence of fluoro moieties which stabilize

monofunctional complex-DNA adducts by their strong hydrogen bonding formation

aptitude (Figs. 3.10 & 3.11) with DNA bases. These hydrogen bonds stabilizing the

monofunctional complex-DNA adducts, are expected to efficiently distort DNA double

helix, and untimely may block transcription. Moreover, IC50 values are much closer to

DNA-distorting anticancer drug cisplatin [14-16] instead of kinase inhibitor

staurosporine, thus revealing the DNA-distortion as a reason for anticancer action. For

the most active complex 1 activity varies against different cell lines in the sequence LU

IcIc7 PC3 MDA-231 MCF7.

Figure 3.9: Comparison of IC50 (µM) values of 1, 2 and 6 against five different cancer

cell lines.

Table 3.12: IC50 values (µM) of the complexes (1, 2 and 6) against five different

cancer cell lines.

3.7

49.3

20.1

112.6

0.03

14.2

45.2

4.9 0.02

4.8

48.4

18.8

38

0.02

6.2

30.1

21.2

0.01

8.8

45.6

22.3

39

0.03 0

20

40

60

80

100

120

1 2 6 Cisplatin Staurosporine

Lu

MCF-7

Hepa-IcIc-7

PC-3

MDA-231

IC5

0 (

µM

)

Compounds

Complex LU MCF-7 Hepa-1c1c7 PC-3 MDA-MB-231

(1) 3.7±1.5 14.2±2.0 4.8±0.8 6.2±1.0 8.8±2.9

(2) 49.27±0.9 45.2±1.3 48.4±5.3 30.1±1.6 45.6±3.8

(6) 20.12±7.4 4.9±0.7 18.8±3.6 21.2±4.3 22.3±2.0

Staurosporine 0.025 0.02 0.020 0.01 0.03

Cisplatin - 22.4 [14] - 38 [15] 39±5.0 [16]

94

Figure 3.10: Structure of 1 showing to have hydrogen bonding ability due to presence of

flouro moieties.

Figure 3.11: Structure of 6 showing to have hydrogen bonding ability due to presence of

flouro moieties.

3.5.3 Anticancer activity against LU human lung carcinoma, MCF-7 human breast

adenocarcinoma

About all the complexes were examined for their in vitro cytotoxic potential against LU

human lung carcinoma and MCF-7 human breast adenocarcinoma by MTT method using

cisplatin as reference drug. The IC50 values are shown in the Table 3.13. The results

95

show that all of these complexes are more active against these two cancer cell lines than

the reference drug cisplatin.

Table 3.13: IC50 values (µM) of the complexes against Lu and MCF-7 cancer cell

lines.

3.6 DNA-Interaction study

DNA is a primary target of the platinum drugs in cancer chemotherapy. Cisplatin forms

mostly intra-strand (96%) cross links among guanine bases after the removal of its two

Complex LU MCF-7 Complex LU MCF-7

(3) 4.91 0.78 (24) 3.36 0.58

(4) 7.48 1.37 (25) 6.89 0.10

(5) 4.33 0.65 (26) 3.69 0.53

(7) 2.94 1.06 (27) 3.69 1.02

(8) 6.56 0.79 (28) 2.20 0.04

(9) 2.40 1.14 (29) - 1.00

(10) 4.60 0.33 (30) 1.57 0.56

(11) 5.95 0.46 (31) 2.37 0.86

(12) 11.04 0.07 (32) 7.61 2.92

(13) 4.91 0.04 (33) 0.99 0.99

(14) 0.93 0.87 (34) 1.20 1.20

(15) 3.35 0.71 (35) 2.95 1.48

(16) 3.62 0.94 (36) 8.99 0.51

(17) 9.12 1.01 (37) 6.86 3.77

(18) 5.51 0.49 (38) - 0.76

(19) 4.15 0.53 (39) 6.44 0.58

(20) 9.97 0.04 (40) 0.80 0.58

(21) 6.24 2.74 (41) 1.33 1.40

(22) 1.12 0.14 (42) 1.67 1.02

(23) 5.13 1.08 cisplatin 16.00 5

96

chloride ligands [17, 18], thus arresting the cellular processes like transcription,

replication and consequently the cell division. Therefore, interaction of new chemical

compounds with DNA becomes one of the most important aspects of biological

investigation in drug development processes. Experimental studies play a basic role for

the understanding of compound-DNA interaction, and provide necessary information for

the predictions of interaction mode and structural analysis. In this work, complex-DNA

interaction has been investigated by UV-Visible spectroscopy and the proposed mode of

interaction has been substantiated by viscometry and cyclic voltammetry.

3.6.1 UV-Visible spectroscopy

UV-Visible spectroscopy is mostly used method for the study of binding constants and

binding modes of metal complexes with DNA. Hypochromism and hyperchromism are

the spectroscopic responses that can be observed due to metal complex-DNA binding,

depending upon the type of interaction. It has been investigated that hypochromism being

the result of - stacking is observed in the case of intercalative binding mode [19].

Absorption spectra of the representative complexes 1, 2, 6, 16, 18 and 19 in the absence

and presence of increasing concentrations of the CT-DNA (at constant concentration of

the complexes) are shown in the Figs. 3.12-3.17. In the figures, low intensity spectra

represent electronic absorptions of the free metal complexes i.e. in the absence of DNA.

The intense intra-ligand π-π* UV region peaks [20] observed in the spectra undergone the

obvious hyperchromic changes upon addition of the increasing DNA concentrations.

These results suggest complex-DNA interaction mode other than intercalation [21]. This

kind of hyperchromic effect can be assigned to electrostatic interaction, a mode

accompanied with distortion in the metal coordination core and as a result enhancing the

probability of intra-ligand π-π* transitions [22]. The electrostatic binding mode shows the

generation of a cationic complex in solution via detachment of the chloride from

platinum centre. Substitution of the chloride with solvent molecules i.e. H2O within the

cell, and then target the DNA is also well recognized [23]. The association/binding

constants were calculated from the intercept-to-slope ratios of Ao/(A -Ao) vs. 1/[DNA]

plots to elucidate the binding strength of these complexes with DNA, and are shown in

Table 3.14. Gibb’s free energies were calculated to explain the spontaneous or non-

97

spontaneous nature of the complex-DNA interactions and are given in Table 3.14.

Negative values of Gibb’s free energy recommend the spontaneous nature of the

complex–DNA interactions.

Table 3.14: Binding constants (K) and Gibb’s free energies (ΔG) of selected Pt(II)

complexes based upon UV- visible spectroscopic data.

Compounds # Binding constant (K) 103 M

-1 Gibb’s free energ (ΔG ) kJ.mol

-1

(1) 18.1 -24.29

(2) 7.40 -22.07

(3) 3.63 -20.31

(5) 1.94 -18.76

(6) 11.2 -23.10

(7) 9.61 -22.72

(8) 4.21 -20.68

(9) 6.51 -21.76

(10) 1.66 -18.37

(16) 8.22 -22.33

(17) 3.75 -20.39

(18) 6.60 -21.79

(19) 2.31 -19.19

98

Figure 3.12: Absorbance of 35 µM complex (1) in the absence (a) and presence of (b) 5

µM, (c) 15 µM, (d) 20 µM, (e) 25 µM and (f) 30 µM DNA. The inset graph

represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for calculation of

binding constant (K) and Gibb’s free energy (ΔG).

99


µM, (c) 10 µM, (d) 15 µM, (e) 20 µM and (f) 25 µM DNA. The inset graph


for calculation of


100


µM, (c) 6 µM, (d) 9 µM, (e) 12 µM, (f) 15 µM and (f) 18 µM DNA. The

inset graph represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for

calculation of binding constant (K) and Gibb’s free energy (ΔG).

101


µM, (c) 6 µM, (d) 9 µM, (e) 12 µM, (f) 15 µM, (g) 18 µM and (h) 21 µM

DNA. The inset graph represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for calculation of binding constant (K) and Gibb’s free energy (ΔG).

300 350 400 450 500 550

0.0

0.2

0.4

0.6

0.8A

bso

rba

nce

Wavelength (nm)

a

h

y = 94.583x + 0.7774

R² = 0.9988

0

5

10

15

20

25

30

35

0 0.1 0.2 0.3 0.4

Ao/A-Ao

1/[DNA]

102

Figure 3.16: Absorbance of 100 µM complex (18) in the absence (a) and presence of (b)

5 µM, (c) 10 µM, (d) 15 µM and (e) 20 µM DNA. The inset graph


for calculation of


300 330 360 390 420 450 480

0.0

0.2

0.4

0.6

0.8

1.0

1.2

Ab

so

rba

nce

Wavelength (nm)

a

e y = 142.66x + 0.9464

R² = 0.9997

0

5

10

15

20

25

30

35

0 0.05 0.1 0.15 0.2 0.25

Ao/A-Ao

1/[DNA]

103

Figure 3.17: Absorbance of 100 µM complex (19) in the absence (a) and presence of (b)

5 µM, (c) 10 µM, (d) 15 µM, (e) 20 µM, (f) 25 µM and (g) 30 µM DNA.

The inset graph represents the plot of Ao/A-Ao vs. 1/[DNA] (µM)-1

for

calculation of binding constant (K) and Gibb’s free energy (ΔG).

3.6.2 Viscometry

Viscosity measurement is a hydrodynamic method to evaluate drug-DNA interaction and

is regarded as the most effective means of distinguishing intercalation and non-

intercalation modes in the absence of crystallographic or NMR data [24]. Effect of

increased complex concentrations on the relative viscosity of CT-DNA for the

representative complexes is shown in Figs. 3.18-3.21. The results, showing decrease in

relative viscosity of CT-DNA with increasing complex concentration, which means

complexes bend/kink the DNA helical structure decreasing its effective length, and hence

the viscosity [24, 25]. These results can be rationalized in terms of non-intercalative

binding, mainly governed by the metal center instead of coordinated ligands [26, 27].

Since the metal complex-DNA binding via electrostatic interaction usually display

300 350 400 450 500

0.0

0.2

0.4

0.6

0.8

1.0

Ab

so

rba

nce

Wavelength (nm)

a

g

y = 160.65x + 0.3709

R² = 0.9971

0

5

10

15

20

25

30

35

0 0.05 0.1 0.15 0.2 0.251/[DNA]

Ao/A-Ao

104

decrease in viscosity of DNA [28], hence herein, is a clue favoring the mode as proposed

via electronic absorption measurements.

0.0 0.5 1.0 1.5 2.0 2.5

0.5

0.6

0.7

0.8

0.9

1.0(

/)1

/3

r

1

2

3

4

5

Figure 3.18: Effects of increasing amounts of complexes 1-5 on the relative viscosities of

CT-DNA at room temperature, [DNA] = 100 µM, r = [complex]/[DNA].

0.0 0.5 1.0 1.5 2.0 2.5

0.5

0.6

0.7

0.8

0.9

1.0

(/

)1

/3

r

6

7

8

9

10

Figure 3.19: Effects of increasing amounts of complexes 6-10 on the relative viscosities

of CT-DNA at room temperature, [DNA] = 100 µM, r = [complex]/[DNA].

105

0.0 0.5 1.0 1.5 2.0 2.5

0.5

0.6

0.7

0.8

0.9

1.0

(/

)1

/3

r

11

12

13

14

15



0.0 0.5 1.0 1.5 2.0 2.5

0.5

0.6

0.7

0.8

0.9

1.0

(/

)1

/3

r

16

17

18

19



106

3.6.3 Cyclic voltammetry

In order to further confirm the complex-DNA interaction a representative complex i.e. 2

was also studied by cyclic voltammetric technique. The complex provides couple of well-

defined redox peaks at 50 mV/s scan rate with an oxidation maximum at -0.6655V and a

reduction maximum at -0.7985V, suggesting a reversible process (Fig. 3.22). Change in

the oxidation peak current was examined after adding the DNA solution. The decrease in

oxidation current from 2.57 10-3

to 1.72 10-3

mA in the presence of 80 µM DNA

signifies complex-DNA interaction (Fig. 3.22). The association/binding constant

calculated (0.74 4 M

-1) from changes in the oxidation peak current by the successive

addition of different DNA concentrations (Figs. 3.23 and 3.24) is in agreement with K

value obtained from electronic absorption data. The lower diffusion coefficients upon 20

µM DNA addition (5.07 10-7

) than the free-complex (5. 47 10-7

) reveal the formation

of high molecular weight complex-DNA adduct (Figs. 3.25-3.27).

Figure 3.22: Representative cyclic voltammogram of 1mM complex 2 in the absence of

DNA (red) and in the presence of 80 µM DNA (black) in DMSO with 0.5

M TBAP as supporting electrolyte at 50 mVs-1

scan rate.

107

Figure 3.23: Cyclic voltamograms of 1 mM complex 2 with 0.5 M TBAP as supporting

electrolyte in the absence (red) and presence of 20 μM DNA (green), 40

μM DNA (black), 60 μM DNA (blue) and 80 μM DNA (orange) showing a

decrease in current.

Figure 3.24: Representative plot of log (I/Io-I) versus log (1/[DNA]) for determination of

binding constant of complex (2).

108

Figure 3.25: Representative cyclic voltammogram of 1 mM complex 2 at different (50-

500 mVs-1

) scan rates in DMSO with 0.5 M TBAP as supporting

electrolyte.

Figure 3.26: Representative cyclic voltammogram of 1 mM complex 2 in the presence

of 20 µM DNA at different (50-500 mVs-1

) scan rates in DMSO with 0.5

M TBAP as supporting electrolyte.

109

Figure 3.27: Representative plot for determination of diffusion coefficients of free drug

(2) and drug-DNA adduct.

3.7 Plasmid DNA cleavage

Effect of the representative complexes (50 µg/mL) on the pUC 19 DNA (12.5 µg/mL) in

tris-HCl buffer (0.1M, pH 7.4) at 37 after incubating for 6 hours was studied using

agarose gel-electrophoresis. Plasmid DNA cleavage by the representative complexes (1,

2, 4-14, 18, 19 and 28) and cisplatin are shown in Figs. 3.28 & 3.29. The

electrophoretograms show that the complexes and cisplatin cleave the DNA, as they

convert supercoiled circular DNA to nicked circular DNA, and the activity markedly

depends on slight variation in the structure. Intensity of the electrophoretogram bands

show that the complexes 1, 4, 7-9, 11-13 and 18 have remarkable DNA cleavage ability,

with the highest one exhibited by 4. The complexes 1 and 4, with the highest DNA

cleavage ability were investigated at different concentrations i.e. 25, 50, 75 and 100

µg/mL. Herein, electrophoretograms (Figs. 3.30a, 3.30b) indicate that the DNA cleavage

ability of the complexes increase with concentration.

110

Figure 3.28: Effect of complexes 4, 5, 7, 8, 9, 10, 11, 12, 13, 14, 18, 19 and 28 (50

µg/ml) on the cleavage of pUC19 DNA (12.5 μg/ml) in Tris–HCl buffer

(0.1 M, pH 7.4) at 37 °C after incubation for 6 h. Form I = suppercoiled

DNA, Form II = nicked circular DNA.

Figure 3.29: Effect of complexes 1, 2 and 6 (50 µg/ml) on the cleavage of pUC19

DNA(12.5 μg/ml) in Tris–HCl buffer (0.1 M, pH 7.4) at 37 °C after

incubation for 6 h. Form I = suppercoiled DNA, Form II = nicked circular

DNA.

111

(a)

(b)

Figure 3.30: Effect of concentration of the complexes (a) 1 and (b) 4, on the cleavage of

pUC19 DNA (12.5 μg/ml) in Tris–HCl buffer (0.1 M, pH 7.4) at 37 °C

after incubation for 6 h.

3.8 Chloride exchange

As DNA binding studies revealed that the complexes interact with DNA via chloride

dissociation generating a cationic complex for the electrostatic interaction. For validation

of the labile nature of the chloride ligand, representative complexes 29, 32, 34 and 35

were reacted with sodium thiocyanate. Appearance of new peaks in IR spectra of the

products i.e. 2055, 2113 cm-1

(29a), 2050, 2113 cm-1

(32a), 2052, 2108 cm-1

(34a) and

2053, 2114 cm-1

(35a), lacking in the corresponding complexes indicate thiocyanate/iso-

thiocyanate attachment to form two linkage isomeric products (Fig. 3.31) [29]. Identical

1H-NMR spectra of the complexes and their corresponding products indicate the

intactness of dithiocarbamate and organophosphine and only chloride replacement by

thiocyanate (Fig. 3.32). Similarly, small up-field shift in 31

P-NMR spectra of the products

compared to the complexes further rectify the assumption (Fig. 3.33).

112

Figure 3.31: Comparison of IR spectra of representative complex 32 and its product 32a

showing appearance of new peaks due to thiocyanate attachment via N or S

atom.

Figure 3.32: Comparison of

1H NMR spectra of representative complex 32 and its

product 32a showing no change in the peak positions.

32

32 a

113

Figure 3.33: Comparison of 31

P NMR spectra of the representative complex 32 and its

product 32a showing small upfield shift.

The chloride substitution was also examined by thiocyanate, bromide, iodide, thiourea

and diethyldithiocarbamate using 31

P-NMR spectroscopy (Table 3.15). In the case of

smaller ions like thiocyanate, bromide and iodide, the chloride replacement was too fast

accompanied by a small 31

P up-field shift. For example, the case of complex 32 in the

presence of equal molar solution of iodide ions; where 31

P-NMR spectra recorded after

the mixing time of 10, 20 and 30 min. indicate quick substitution of chloride (Fig. 3.34).

Contrarily, the reactions with bulky thiourea (34) and diethyldithiocarbamate (32) were

slower as signified by their 31

P-NMR spectra (Fig. 3.35 and 3.36). Furthermore, two

products were formed as the reaction proceeds in the latter case (Fig. 3.36).

32 32 a

114

Figure 3.34: 31

P NMR spectra of the complex 32 (25mM) in the presence of equal molar

solution of iodide ions recorded after 10, 20 and 30 min. of mixing.

Figure 3.35:

31P NMR spectra of the complex 34 (25 mM) in the presence of excess of

thiourea recorded after 2, 4 and 6 h of mixing.

(32)

(32) in the presence of equal molar I- ion

after 10, 20 and 30 min of mixing

115

Figure 3.36:

31P NMR spectra of the complex 32 (10 mM) in the presence of equal molar

solution of diethyldithiocarbamate recorded after different intervals of time.

Table 3.15: 31

P NMR peaks (ppm) of the representative complexes in the presence of

different exchangeable ligands.

Complex Exchangeable

ligand

31P NMR

(ppm)

(18) Cl 13.31

I 13.25

(29) Cl 13.23

SCN 13.19

Br 13.17

I 13.12

(32) Cl 13.33

SCN 13.29

Br 13.30

I 13.28

DT 13.20, 13.37

(34) Cl 14.42

SCN 14.37

Br 14.38

I 14.35

Thu 14.38

(35) Cl 11.30

SCN 11.24

Br 11.26

I 11.23

After 15 min of mixing

After 1 h of mixing

After 2 h of mixing

After 4 h of mixing

After 6 h of mixing

After 8 h of mixing

After 10 h of mixing

After 12 h of mixing

116

Exploiting the slow chloride exchange by diethyldithiocarbamate (in excess) for 32,

kinetics of the reaction under pseudo-first order conditions was investigated by UV-

Visible spectroscopy. The change in absorbance with time indicated the reaction

progress. The apparent rate constant was calculated from the straight-line slope, obtained

from lnΔA vs t. Also, different apparent rate constants were determined with different

diethyldithiocarbamate concentrations (Figs. 3.37-3.40). When these apparent rate

constants were plotted against different diethyldithiocarbamate concentrations, a straight

line was obtained (Fig. 3.41) showing first order kinetics of the reaction. Herein, slope of

the plot gives the value of observed rate constant (Fig. 3.41).

Figure 3.37: Spectrum of change of absorbance with respect to time using

diethyldithiocarbamate (110 µM) and complex 32 (10 µM). Inset is the

graphs between ln∆A verses time.

260 280 300 320 3400.0

0.2

0.4

0.6

0.8

1.0

1 2 3 4 5 60

-1

-2

-3

-4

-5

y = -0.2379x - 3.3714

R² = 0.9949

Ab

so

rba

nce

Wavelength (nm)

ln

A

t (min)

117




Figure 3.39: Spectrum of change of absorbance with respect to time using diethyldithioc-

arbamate (120 µM) and complex 32 (10 µM). Inset is the graphs between

ln∆A verses time.

260 280 300 320 3400.0

0.2

0.4

0.6

0.8

1.0

1 2 3 4 5 60

-1

-2

-3

-4

-5

Ab

so

rba

nce

Wavelength (nm)

y = -0.2464x - 2.9158

R² = 0.9937

ln

A

t (min)

260 280 300 320 3400.0

0.2

0.4

0.6

0.8

1.0

1 2 3 4 5 60

-1

-2

-3

-4

-5

y = -0.2521x - 2.9357

R² = 0.993

t (min)

Ab

so

rba

nce

Wavelength (nm)

ln

A

118




Figure 3.41: Graph between apparent rate constants vs different concentrations of DT

(diethyldithiocarbamate).

260 280 300 320 3400.0

0.2

0.4

0.6

0.8

1.0

1.2

1 2 3 4 5 60

-1

-2

-3

-4

-5

Ab

so

rba

nce

Wavelength (nm)

y = -0.2652x - 2.6906

R² = 0.993

ln

A

t (min)

y = 1752x + 0.0445

R² = 0.9734

0.18

0.19

0.2

0.21

0.22

0.23

0.24

0.25

0.26

0.27

1.05 1.1 1.15 1.2 1.25 1.3

[DT] X 10-4M

K a

pp (

min

-1)

119

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Navarro, M., Chiral platinum(II) complexes featuring phosphine and chloroquine

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Chapter 4

CRYSTALLOGRAPHIC ANALYSIS AND DFT STUDIES

123

4.1 Single crystal X-ray analysis

Crystal data and structure refinement parameters for the heteroleptic Pt(II) complexes 1,

2, 6, 25 and 29 with different crystal systems (monoclinic & orthorhombic) and space

groups (P21/c & Pbca) are given in the Tables 4.1 & 4.2. The selected bond lengths (Å)

and bond angles (˚) are shown in the Tables 4.3 & 4.4 respectively.

Monofunctional complexes (1, 2, 6, 25) show pseudo square-planar geometry around

platinum atom with two cis sites occupied by dithiocarbamate chelate forming four-

membered ring (PtS2C), along with cis positioned chloride and organophosphine (Figs.

4.1 and 4.2). The S(1)-Pt(1)-S(2) angles {74.91(4 1, 2, 75.04(6 6, 75.31(3

25} are smaller than ideal value 90 confirming pseudo cis position of the two sulfur

atoms due to chelate restriction. This distortion also lowers down trans S(1)-Pt(1)-Cl(1)

angle {170.37(4 1, 168.52(1 2 and 168.16(6 6, 169.42(3 25} by around 10-12o than

ideal value of 180 , thus resulting in pseudo square planar geometry. Two sulfurs

coordinate slightly asymmetrically with a short {2.2766(12) 1; 2.293(4) 2; 2.2923(16) 6,

2.2821(8) 25} and a long {2.3574(12) 1; 2.343(4) 2; 2.3527(15) 6, 2.3556(9) 25} Pt-S

bond being attributed to the trans-influence of the phosphine. However, the magnitude of

asymmetry varies in these complexes {ΔPt-S of 0.0808 Å (1) 0.05 Å (2) 0.0604 Å (6),

0.0.0735 Å 25}. This asymmetry in Pt-S bonds due to trans effect of the ligand is typical

for square-planar system. The bond lengths of C(1)-S(1) {1.718(4) Å 1, 1.741(18) Å 2,

1.728(6) Å 6, 1.730(3) Å 25} and C(1)-S(2) {1.717(5) Å 1, 1.712(14) 2, 1.724(6)Å 6,

1.721(3) Å 25} are in between C-S single (1.82 Å) and double bond (1.60 Å) [1].

Similarly, C(1)-N(1) bond lengths 1.306(5) Å 1,1.31(2) Å 2 and 1.314(7) Å 6, 1.309(4) Å

25} are shorter than single (1.47 Å) and longer than double bond (1.28 Å) [2], thus

confirming a resonance phenomenon in NCSS moiety.

Similarly, crystal structures of bis-organophosphine complex (29) having no replaceable

chloride also shows pseudo square-planar geometry around platinum atom with two cis

sites occupied by dithiocarbamate chelate forming four-membered ring (PtS2C). While

two remaining cis sites occupied by two phosphorous atoms of 4-

bis(diphenylphosphino)butane to form seven-membered metallacycle (Fig. 4.2). Due to

124

trans-effect of both phosphorous atoms of bis-phosphine, dithiocarbamate in complex

(29) is loosely attached compared to that in monofunctional complexes (1, 2, 6 and 25) as

indicated by their bond lengths (Table 4.3). Similarly the magnitude of asymmetry is less

in this complex {ΔPt-S of 0.0292 Å (29)} as compared to that in monofunctional

complexes (1, 2, 6 and 25). Additionally, a solvent molecule i.e. MeOH, is also present in

the single crystal XRD structure of the complex.

Figure 4.1: Structures of monofunctional complexes 1, 2, 6 and 25.

Figure 4.2: Structure of complex 29 showing pseudo square planar geometry.

125

Table 4.1: Crystal data and structure refinement for the complexes 1, 2 and 6.

Crystal Data 1 2 6

Chemical formula C30H27ClF3N2OPPtS2 C30H27Cl4N2OPPtS2 C28H23ClF3N2O2PPtS2

Crystal size

(mm3)

0.350 × 0.220 × 0.160 0.400 × 0.230 × 0.160 0.38 × 0.20 × 0.15

Formula weight

(g mol-1

)

814.16 863.51 802.11

Crystal system Monoclinic Monoclinic Orthorhombic

Space group P21/c P21/c Pbca

Temperature (K) 296(2) 296(2) 296(2)

Radiation MoKα λ = 0 0 3 MoKα λ = 0 0 3 MoKα λ = 0 0 3

Cell parameters

a (Å) 19.2669(11) 20.5863(15) 9.4060(4)

b (Å) 9.8920(6) 9.3450(7) 17.1884(10)

c (Å) 16.4584(9) 18.4610(13) 36.1133(19)

α ° 90 90 90

β ° 96.837(3) 114.952(3) 90

γ ° 90 90 90

Volume (Å3) 3114.5(3) 3220.0(4) 5838.6(5)

Z 4 4 8

μ mm-1

) 4.821 4.897 5.145

F(000) 1592.0 1688.0 3120.0

2Θ range for data

Collection

4.636 to 56.034 4.414 to 55.202 4.74 to 55.046

Reflections collected 27678 26012 27915

Independent reflections 7449 7384 6716

Goodness-of-fit on F2 0.976 1.130 1.110

Final R indexes [I>=2σ

(I)]

R1 = 0.0386,

wR2 = 0.0663

R1 = 0.0815,

wR2 = 0.1982

R1 = 0.0458,

wR2 = 0.0770

Final R indexes [all data] R1 = 0.0752,

wR2 = 0.0760

R1 = 0.1195,

wR2 = 0.2127

R1 = 0.0914,

wR2 = 0.0877

126

Table 4.2: Crystal data and structure refinement for the

complexes 25 and 29. Crystal Data 25 29

Chemical formula C30H27ClF3N2PPtS2 C41H47ClN2OP2PtS2

Crystal size (mm3) 0.12 × 0.06 × 0.06 0.16 × 0.12 × 0.1

Formula weight (g mol-1) 798.16 940.40

Crystal system Orthorhombic Monoclinic

Space group Pbca P21/c

Temperature (K) 100 100

Radiation GaKα λ = 3 3 GaKα λ = 3 3

Cell parameters

a (Å) 9.3759(5) 14.5420(6)

b (Å) 16.9882(8) 9.5405(4)

c (Å) 37.4297(18) 28.1731(11)

α ° 90 90

β ° 90 92.3180(10)

γ ° 90 90

Volume (Å3) 5961.8(5) 3905.5(3)

Z 8 4

μ mm-1) 8.210 6.514

F(000) 3120.0 1888.0

2Θ range for data

collection

8.22 to 121.574 5.292 to 114.006

Reflections collected 51066 61767

Independent reflections 6817 7971

Goodness-of-fit on F2 1.119 1.080

Final R indexes [I>=2σ I ] R1 = 0.0307,

wR2 = 0.0710

R1 = 0.0176,

wR2 = 0.0435

Final R indexes [all data] R1 = 0.0368,

wR2 = 0.0736

R1 = 0.0182,

wR2 = 0.0438

127

It is known that square planar platinum(II) complexes undergo ligand substitution

reactions by an associative mechanism at axial position expectedly through trigonal

bipyramidal transition state. In monofunctional complexes (1, 2, 6 and 25),

organophosphine moiety is oriented such that two of its aromatic rings are placed

approximately perpendicular to coordination plane, while the third one is in plane.

Consequently, one of aromatic C-H occupies approximately axial position of platinum

coordination plane (Fig. 4.3), thus causing steric hindrance for axially approaching

nucleophile, a phenomenon also observed in picoplatin and phenanthriplatin [3-4].

Table 4.3: Selected bond lengths of the complexes 1, 2, 6, 25 and 29.

Bond Type

Bond lengths(Å)

1 2 6 25 29

Pt1-P1 2.2361(12) 2.231(3) 2.2482(15) 2.2396(9) 2.2735(4)

Pt1-P2 - - - - 2.2835(4)

Pt1-S1 2.2766(12) 2.293(4) 2.2923(16) 2.2821(8) 2.3477(4)

Pt1-S2 2.3574(12) 2.343(4) 2.3527(15) 2.3556(9) 2.3769(4)

Pt1-Cl1 2.3340(12) 2.334(4) 2.3283(16) 2.3368(8) -

S1-C1 1.718(4) 1.741(18) 1.728(6) 1.730(3) 1.7273(18)

S2-C1 1.717(5) 1.712(14) 1.724(6) 1.721(3) 1.7318(19)

N1-C1 1.306(5) 1.31(2) 1.314(7) 1.309(4) 1.317(2)

Table 4.4: Selected bond angles of the complexes 1, 2, 6, 25 and 29.

Angle type Bond angle (˚)

1 2 6 25 29

P1-Pt1-S1 96.37(4) 97.98(12) 97.61(6) 96.75(3) 94.095(16)

P1-Pt1-Cl1 93.16(5) 92.47(15) 94.03(6) 93.61(3) -

S1-Pt1-Cl1 170.37(4) 168.52(15) 168.16(6) 169.42(3) -

P1-Pt1-S2 170.23(4) 172.50(13) 171.00(5) 171.29(3) 168.272(16)

S1-Pt1-S2 74.91(4) 74.97(14) 75.04(6) 75.31(3) 74.302(15)

Cl1-Pt1-S2 95.69(4) 94.78(17) 93.56(6) 94.49(3) -

P1-Pt1-P2 - - - - 94.222(16)

P2-Pt1-S1 - - - - 171.596(15)

P2-Pt1-S2 - - - - 97.346(15)

128

Complexes 1 and 25 are protected at both axial sites by ArC-H, contrary to other two

complexes (2 & 6) where only one side is shielded. The distance of C-atoms (attached to

shielding H) from Pt on both axial sites is unidentical (i.e. 3.435 Å and 3.635 Å)

indicating relatively more hindrance at one side in 1. In 25 this distance is almost

identical (i.e. 3.594 Å and 3.634 Å) indicating equal hindrance at both axial sides.

However, these distances are slightly longer than observed for picoplatin (3.224 Å) and

phenanthriplatin (3.220 Å) [3]. In other complexes the Pt···C distances are 3.616 (2) and

3.670 Å (6), along with insignificant complementary axial hindrance due to the ring

orientations. Thus, complex 1 is expected to be more inert for substitution reactions

because of greater and dual steric hindrance.

4.2 Crystal packing

These complexes provide an ideal platform to perceive the effect of various weak CH···X

[X = O, F, Cl and π] non-covalent interactions on the solid-state self-assembly

particularly in the absence of strong hydrogen bonds [5-14]. The crystal structure of

monofunctional complexes (1, 2, 6 and 25) revealed 3D-networks, composed of various

1D-supramolecular chains stabilized by variety of weak non-covalent interactions. The

molecules are arranged in head to head fashion in 1D-chains of compound 1 stabilized by

C-H···F [(C(18)-H(18)···F(2A) 2.529 Å, (C(17)-H(17)···F(2A) 2.648 Å, (C(30)-

H(30)···F(2A) 2.622 Å], C-H···O [(C(15)-H(15)···O(1) 2.705 Å] and C-H···π [(C(15)-

H(15)···C(9) 2.884 Å, (C(14)-H(14)···C(10) 2.825 Å] interactions (Fig. 4.4a, Table 4.5),

antiparallel fashion in compound 2 stabilized by C-H···Cl [(C(18)-H(18)···Cl(4) 2.889 Å,

(C(11)-H(11)···Cl(1) 2.768 Å] and C-H···π [(C(23)-H(23)···C(10) 2.882 Å, (C(23)-

H(23)···C(11) 2.881 Å] interactions (Fig. 4.4b, Table 4.5), slightly displaced antiparallel

fashion in compound 6 stabilized by C-H···F [(C(24)-H(24)···F(2) 2.434 Å], C-H···S

[(C(9)-H(9)···S(1) 2.901 Å] and C-H···π [(C(9)-H(9)···C(1) 2.876 Å, (C(10)-

H(10)···C(18) 2.846 Å] interactions (Fig. 4.4c, Table 4.5) and parallel fashion in

compound 25 stabilized by C-H···Cl [(C(2)-H(2A)···Cl(1) 2.889 Å], C-H···S [(C(3)-

H(3A)···S(2) 2.975 Å], C-H···π [(C(5)-H(5B)···C(11) 2.844 Å], π···π [(C(5)···C(11)

3.341 Å] and F···π [(F(3)···C(15) 2.993 Å] interactions (Fig. 4.4d, Table 4.5).

129

The 1D-supramolecules extend themselves by means of C-H···Cl [(C(5)-H(5B)···Cl(1)

3.017 Å], C-H···S [ (C(4)-H(4B)···S(2) 3.068 Å], C-H···O [ (C(2)-H(2B)···O(1) 2.814

Å] and C-H···π [(C(2)-H(2A)···C(9) 2.935 Å] interactions in compound 1 (Fig. 4.5a), by

means of C-H···Cl [(C(29)-H(29)···Cl(2) 3.262 Å, (C(3)-H(3A)···Cl(1) 3.017 Å],

halide···π [Cl(2)···C(29) 3.436 Å] and C-H···π [(C(15)-H(15)···C(21) 2.848 Å, (C(15)-

H(15)···C(22)2.963 Å] interactions in compound 2 (Fig. 4.5b), by means of C-H···O

[(C(19)-H(19)···O(1) 2.631 Å, (C(18)-H(18)···O(1) 2.708 Å, (C(13)-H(13)···O(1) 2.668

Å], C-H···S [(C(3)-H(3B)···S(2) 3.068 Å], C-H···π [(C(5)-H(5A)···C(8) 2.996 Å, (C(5)-

H(5A)···C(10) 2.768 Å, (C(13)-H(13)···C(8) 2.911 Å] interactions in compound 6 (Fig.

4.5c, Table 4.5) and by means of C-H···S [(C(9)-H(9B)···S(1) 2.877 Å, (C(2)-

H(2B)···S(2) 2.979 Å], C-H···F [(C(20)-H(20)···F(1) 2.508 Å, (C(26)-H(26)···F(1)

2.367 Å, (C(21)-H(21)···F(3) 2.453 Å,], F···π [(F(1)···C(26) 3.080 Å] interactions in

compound 25 (Fig. 4.5d, Table 4.5) to provide an overall 3D-network structures. These

results indicate the importance of weak non-covalent interactions for the formation of

1D-network and overall 3D-network structures. The findings also suggest the strong

ability of the complexes to interact with surroundings through these interactions which

are expected to play very important role not only in solid state but also in solution. In the

complex 29 1D-supramolecular chain is formed by methanol bridging between piperazine

ring of dithiocarbamate and aromatic ring of organophosphine by C-H···O [(C(3)-

H(3B)···O(1) 2.529 Å, (C(38)-H(38)···O(1) 2.648 Å], C-H···H [(C(38)-H(38)···H(1)

2.705 Å] and C-H···C [(C(38)-H(38)···C(41) 2.884 Å] interactions (Fig. 4.6). The 1D-

supramolecular chain extend itself in three dimensions by means of C-H···Cl [(C(5)-

H(5B)···Cl(1) 3.017 Å], C-H···S [(C(4)-H(4B)···S(2) 3.068 Å], C-H···O [(C(2)-

H(2B)···O(1) 2.814 Å] and C-H···π [(C(2)-H(2A)···C(9) 2.935 Å] interactions.

130

Figure 4.3: Crystal structures of the complexes (1, 2, 6 and 25) showing steric hindrance

from aromatic C-H groups of organophosphine, C-Pt distance (black), H-Pt

distance (red).

3.634Å

3.594Å

3.135Å

3.059Å

(25)

3.67Å 3.16Å

(6)

3.635Å

3.435Å

3.076Å

2.851Å

3.616Å

3.112Å

(1) (2)

131

(a) (b)

(c) (d)

Figure 4.4: 1D-supramolecular chains in 3D-crystal packing of 1, 2, 6 and 25.

132

(a) (b)

(c)

(d)

Figure 4.5: The 3D-crystal packing of 1 along b-axis (a), 2 along b-axis (b), 6 along a-

axis (c) and 25 along b-axis (d).

133

X= F (1, 6, 25), Cl (2)

Table 4.5: Various weak non-covalent interactions (Å) forming 1D and 3D networks for crystal packing. Complex C-H···X C-H···O C-H···π C-H···S C-H···Cl X···π

(1)

1D 3D 1D 3D 1D 3D

1D 3D 1D 3D 1D 3D

2.529

2.648

2.622

-

2.705

2.814

2.884

2.825

2.935

-

3.068

-

3.017

-

-

(2)

2.889

2.768

3.262

-

-

2.882

2.881

2.848

2.963

-

-

-

3.017

-

3.436

(6)

2.434

-

-

2.631

2.708

2.668

2.876

2.846

2.996

2.768

2.911

2.901

3.068

-

-

-

-

(25) 2.508

2.367

2.453

2.844 2.975 2.877

2.979

2.889 2.993 3.080

134

(a) (b)

Figure 4.6: The crystal packing of 29, 1D-packing through methanol bridging (a), 3D-

packing along a-axis (b).

4.3 Theoretical studies

Theoretical calculations were performed to assess the properties and reactivity of the

synthesized complexes. The geometry optimization was performed at DFT/B3LYP level

of the theory using LANL2DZ basis set [15-18] to obtain the lowest energy structure of

the complexes. DFT based optimized structures of the monofunctional complexes are

almost identical to that obtained from single crystal XRD (Fig. 4.7, Table 4.6 & 4.7). All

of the complexes adopt pseudo square planar geometry around platinum atom with cis S-

Pt-S and trans S-Pt-Cl angles smaller than ideal 0˚ and 80˚ respectively, due to four-

membered PtS2C chelate restriction. However, theoretical bond lengths are slightly

greater than the experimental values (Table 4.6 & 4.7). Likewise, the results indicate that

the pronounced asymmetric behavior of the Pt-S distances [Pt1-S1 = 2.2766 Å, Pt1-S2 =

2.3574 Å (1), Pt1-S1 = 2.293 Å, Pt1-S2 = 2.343 Å (2), Pt1-S1 = 2.2923 Å, Pt1-S2 =

135

2.3527 Å (6) and Pt1-S1 = 2.2821 Å, Pt1-S2 = 2.3556 Å (25)] observed in the single cry-

stal structures, is absent in the optimized structures [Pt1-S1 = 2.4559 Å, Pt1-S2 = 2.4771

Å in (1), Pt1-S1 = 2.4556 Å, Pt1-S2 = 2.4767 Å in (2), Pt1-S1 = 2.456 Å, Pt1-S2 =

2.4787 Å in (6) and (Pt1-S1 = 2.4533 Å, Pt1-S2 = 2.475 Å in (25)]. The difference of the

theoretical and experimental data can be attributed to the gas phase optimization, that is

obviously, deprived of bulk interactions [19].

Like the XRD single crystals, axial protection of ArC-H of the organophosphine is also

observed in the DFT-optimized structures (Fig. 4.8). Spatial orientation of the organo-

phosphine renders this steric protection to platinum from axial off-target attacks.

Complexes 1 and 25 are sterically protected at both, while all of the others at single axial

site. Theoretically calculated Pt···C and Pt···H distances (of shielding H and C-atoms) of

some representative monofunctional complexes are collected in the Table 4.8. It can be

noticed that the distances obtained from optimized structures are slightly larger than that

of single crystal XRD structures. These slight differences can be correlated with the

unavailability of secondary forces in the gas phase optimization.

(a) (b)

Figure 4.7: Comparison of single crystal XRD structure (a) and optimized structure (b)

of the representative complex 2.

Molecular properties like dipole moment, molecular polarizability and electronic

structure etc. are affected by the charge on each atom of the compound. Moreover,

136

measurement of atomic charges of the compounds is also important in assessing their

reactivity. Therefore, estimation of the Mullikan atomic charges becomes important in

quantum chemical calculations. To this end, natural population analysis reveals improved

numerical stability and better depiction of electronic distribution in the compounds of

high ionic character like those containing metal atoms [20]. Natural population analysis

was carried out to assess NBO atomic charges after DFT optimization and full NBO run,

using same method and same basis set [14-18]. NBO atomic charges via color schemes

Table 4.6: Comparison of the selected bond lengths by theoretical and X-ray calculations for

the complexes 1, 2, 6 and 25. Type of

Bond

Bond lengths(Å) from single crystal XRD

Bond lengths(Å) from DFT/LANL2DZ

1 2 6 25 1 2 6 25

Pt1-P1 2.2361(12) 2.231(3) 2.2482(15) 2.2396(9) 2.3652 2.3649 2.3664 2.3674

Pt1-S1 2.2766(12) 2.293(4) 2.2923(16) 2.2821(8) 2.4559 2.4556 2.456 2.4533

Pt1-S2 2.3574(12) 2.343(4) 2.3527(15) 2.3556(9) 2.4771 2.4767 2.4787 2.475

Pt1-Cl1 2.3340(12) 2.334(4) 2.3283(16) 2.3368(8) 2.4368 2.4367 2.4353 2.4378

S1-C1 1.718(4) 1.741(18) 1.728(6) 1.730(3) 1.7987 1.7986 1.7969 1.8015

S2-C1 1.717(5) 1.712(14) 1.724(6) 1.721(3) 1.7831 1.7834 1.7833 1.7842

N1-C1 1.306(5) 1.31(2) 1.314(7) 1.309(4) 1.342 1.3419 1.3427 1.3409

Table 4.7: Comparison of the selected bond angles by theoretical and X-ray calculations for

the complexes 1, 2, 6 and 25. Angle type

Bond angle (˚) from single crystal XRD

Bond angle (˚) from DFT/LANL2DZ

P1-Pt1-S1 96.37(4) 97.98(12) 97.61(6) 96.75(3) 99.96 99.98 99.96 99.99

P1-Pt1-Cl1 93.16(5) 92.47(15) 94.03(6) 93.61(3) 90.87 90.76 90.91 90.83

S1-Pt1-Cl1 170.37(4) 168.52(15) 168.16(6) 169.42(3) 169.17 169.26 169.13 169.10

P1-Pt1-S2 170.23(4) 172.50(13) 171.00(5) 171.29(3) 174.36 174.38 174.32 174.49

S1-Pt1-S2 74.91(4) 74.97(14) 75.04(6) 75.31(3) 74.40 74.41 74.36 74.42

Cl1-Pt1-S2 95.69(4) 94.78(17) 93.56(6) 94.49(3) 94.76 94.85 94.76 94.68

137

for the representative mono-functional complexes are shown in Fig. 4.9 and the values

are collected in the Table 4.9.

(1) (2)

Figure 4.8: Optimized structures of the complexes (1 and 2) showing steric hindrance

from aromatic C-H groups of organophosphine, C-Pt distance (black), H-Pt

distance (red).

Table 4.8: Pt···H and Pt···C distances (Å) of the optimized structures of the

representative complexes.

Complex Pt···H (Å) Pt···C (Å) Complex Pt···H (Å) Pt···C (Å)

1 3.223

2.997

3.781

3.654

17 3.021 3.665

2 3.006 3.658 18 3.034 3.669

4 3.240

3.037

3.790

3.672

25 3.229

2.986

3.789

3.645

6 2.989 3.643 27 2.866 3.530

7 2.986 3.642 30 2.991 3.649

9 3.031 3.665 31 3.008 3.658

16 2.999 3.653 32 3.030 3.670

Negative atomic charge of the central platinum atom can be rationalized in terms of

electron density shift from the attached organophosphine, dithiocarbamate and chloride

ligands. Donor S and P atoms exhibit positive charges as electron density is transferred to

the central metal atom and form stable Pt-P and Pt-S bonds. Being the stable nature of the

bonds, organophosphine and dithiocarbamate ligands are likely to resist the substitution

138

reactions in biological fluids. Negatively charged Cl¯ ligand is a good leaving entity, as it

possesses the same charge as the central metal atom. In concentrated Cl¯ environments

like extracellular fluids, these complexes are expected to be stable but intracellularly,

Figure 4.9: Structures of representative monofunctional complexes having atomic charge

at each atom representing from color of each atom, assessed from natural

population analysis. Change from green or red to black represents change of

atomic charge (positive or negative) towards neutrality.

where the Cl¯ concentration lowers down, substitution is likely to occur. Besides,

intrinsic polarity of the atoms as evidenced from variations in atomic charges (Fig. 4.9 &

Table 4.9) suggests strong ability of the complexes to involve with surroundings via

secondary interactions.

Molecular properties like dipole moment, frontier molecular orbitals (HOMO & LUMO),

HOMO–LUMO band gap ΔE , ionization potential IP , electron affinity EA , global

hardness η , chemical potential μ and global electrophilicity ω have been assessed

after DFT optimization (Table 4.10) [21-22]. High values for dipole moments represent

polar nature of the complexes, making them liable for secondary interactions.

Frontier molecular orbitals exist at outer edge of a molecule including HOMO and

LUMO. Removal of HOMO electron is the easiest one and negative of its energy is a

good approximation of the experimental ionization energy (IE). High values of IE show

very stable nature of the complexes while, lower LUMO energies illustrate them as soft

acids (SHAB concept) [23]. LUMO can accept electrons and negative of its energy

corresponds to the electron affinity (EA). The calculated EAs for the monofunctional

complexes are positive, showing their potential to react with nucleophiles through LUMO

[24-25]. Furthermore, more than half of the complexes have greater EAs than the isolated

platinum atom (2.125 eV) [26]. Frontier molecular orbitals and their energy difference for

139

the representative monofunctional complexes are shown in Fig. 4.10 & Table 4.10. Large

energy difference is an indication of stable nature of the complexes [27]. Chemical

potential evaluated for the complexes show their ability to undergo spontaneous

reactions. The chemical potential can also be correlated with the structures i.e. complexes

with ArC-H protected platinum center from both the axial sites, have low potential to

undergo spontaneous reactions and vice versa for the one side shielded complexes.

Figure 4.10: Frontier orbitals of the representative complexes showing energy difference

between HOMO and LUMO orbitals.

140

Table 4.9: NBO atomic charges of representative complexes from natural population analysis.

Atoms

NBO atomic charges

1 2 3 4 6 7 8 9 16 17 18

Pt -0.19427 -0.19382 -0.19041 -0.18515 -0.19223 0.19252 -0.19012 -0.18348 -0.18876 -0.18117 -0.18009

Cl -0.39582 -0.39541 -0.39682 -0.39852 -0.39513 -0.39463 -0.39642 -0.39788 -0.38938 -0.39131 -0.39210

P 1.09772 1.09693 1.09943 1.10121 1.09832 1.09693 1.09956 1.10191 1.10088 1.10025 1.10480

S 0.03185 0.03191 0.02963 0.01760 0.02853 0.02809 0.02976 0.01453 0.03490 0.02378 0.02048

S 0.00546 0.00663 0.00572 0.00494 0.01168 0.01422 0.01161 0.01107 0.00875 0.00920 0.00862

C(NCSS) -0.03036 -0.03017 -0.03249 -0.03665 -0.03233 -0.03220 -0.03245 -0.03873 -0.03770 -0.04350 -0.04433

N(NCSS) -0.45444 -0.45424 -0.45768 -0.45935 -0.45708 -0.45693 -0.45747 -0.46191 -0.46015 -0.46441 -0.46515

20 21 22 23 25 26 27 29 30 31 32

Pt -0.19267 -0.19136 -0.19022 -0.18894 -0.196 -0.192 -0.194 -0.19343 -0.19338 -0.18545 -0.18682

Cl -0.39974 -0.39532 -0.39998 -0.39987 -0.398 -0.399 -0.399 -0.39594 -0.39494 -0.39742 -0.39746

P 1.09686 1.09595 1.09889 1.09989 1.096 1.098 1.099 1.09811 1.09719 1.09739 1.09689

S 0.03089 0.03121 0.02843 0.01983 0.033 0.035 0.026 0.03146 0.03169 0.01983 0.01975

S 0.00682 0.00759 0.00498 0.00435 -0.003 -0.003 -0.002 0.00629 0.00720 0.00626 0.00597

C (NCSS) -0.03237 -0.03216 -0.03368 -0.03753 -0.029 -0.031 -0.033 -0.03093 -0.03067 -0.03637 -0.03668

N(NCSS) -0.45557 -0.45533 -0.45868 -0.45998 -0.456 -0.455 -0.448 -0.45491 -0.45469 -0.45912 -0.45162

141

Table 4.10: Molecular properties of representative complexes.

Molecular property Pt(II) complexes

1 2 3 4 6 7 8 9 16 17 18

Dipole moment (Debye) 7.96 6.99 8.03 8.23 5.90 5.79 6.04 4.94 5.10 8.90 9.24

IE = HOMO (eV) 5.29 5.29 5.18 5.15 6.05 6.06 5.94 5.60 6.25 5.87 5.80

EA = LUMO (eV) 2.21 2.23 2.04 1.75 2.25 2.27 2.13 1.79 2.94 2.84 2.83

∆ = (LUMO HOMO) (eV) 3.08 3.05 3.14 3.40 3.80 3.79 3.81 3.81 3.31 3.03 2.97

Global hardness η 1.54 1.53 1.57 1.70 1.90 1.89 1.91 1.91 1.65 1.51 1.48

Chemical potential (µ) -3.75 -3.76 -3.61 -3.45 -4.15 -4.17 -4.04 -3.70 -4.59 -4.35 4.31

20 21 22 24 25 26 27 29 30 31 32

Dipole moment (Debye) 5.67 6.63 7.65 7.92 7.43 8.12 8.84 6.13 6.09 6.52 6.73

IE = HOMO (eV) 5.42 5.39 5.24 5.17 5.95 5.73 5.39 5.40 5.40 5.27 5.23

EA = LUMO (eV) 2.35 2.38 2.10 1.88 2.17 1.97 1.62 2.23 2.25 1.84 1.71

∆ = (LUMO HOMO) (eV) 3.07 3.01 3.14 3.29 3.78 3.76 3.77 3.17 3.15 3.43 3.52

Global hardness η 1.54 1.51 1.57 1.65 1.89 1.88 1.88 1.59 1.58 1.72 1.76

Chemical potential (µ) -3.89 -3.89 -3.67 -3.53 -4.06 -3.85 -3.51 -3.82 -3.83 -3.56 -3.47

142

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145

CONCLUSION

New heteroleptic platinum(II) dithiocarbamates have been synthesized and characterized

by different analytical techniques such as elemental analysis, FT-IR, multinuclear (1H,

13C and

31P) NMR and X-ray single crystal analysis along with DFT calculations. In the

FT-IR spectra, the presence of Pt-S peak (345-390 cm-1

) and an increase in the stretching

frequency for C-N (1479-1540 cm-1

) upon complexation than the free ligands (1433-1474

cm-1

) indicated the platinum-dithiocarbamate coordination. The Pt-Cl, only in mono-

functional complexes, and Pt-P peaks were observed at 273-316 cm-1

and 211-240 cm-1

,

respectively. Additionally, complexes formation was further confirmed by the occurrence

of proton resonances for both dithiocarbamate and organophosphine. In 13

C NMR, an

upfield shift in CSS signal of complexes (197.6-211.0 ppm) than the free ligands (212.7-

214.9 ppm) confirmed the Pt-dithiocarbamate coordination. This upfield shift can be

attributed to shielding of the CSS carbon due to mobilization of the nitrogen lone pair

upon complexation. The organophosphine-platinum coordination was assessed by the

downfield shift in 31

P NMR signal than the free organophosphine. The crystal structures

of monofunctional complexes (1, 2, 6 and 25) showed pseudo square planar geometries

imposed by the chelate restriction of dithiocarbamate that also resulted in Pt-Sshort

{2.2766(12) 1; 2.293(4) 2; 2.2923(16) 6, 2.2821(8) 25} and Pt-Slong {2.3574(12) 1;

2.343(4) 2; 2.3527(15) 6, 2.3556(9) 25} bonds. This dissimilarity in Pt-S bonds can also

be attributed to more tran-effect of the organophosphine than chloride. Due to trans-

effect of both phosphorous atoms of the bis-organophosphine, dithiocarbamate ligand in

complex 29 is relatively loosely attached, as established by Pt-S bond lengths, compare to

that in monofunctional complexes (1, 2, 6 and 25). Similarly, the magnitude of

asymmetry is less in 29 {ΔPt-S of 0.0292 Å} than the monofunctional ones {ΔPt-S of

0.0808 Å 1, 0.05 Å 2 0.0604 Å 6, 0.0.0735 Å 25}. The C-S bond lengths are in between

C-S single (1.82 Å) and double (1.60 Å) bonds. Similarly, C-N bond lengths are shorter

than the single (1.47 Å) and longer than the double (1.28 Å) bonds, thus confirming the

resonance phenomenon in NCSS moiety. The packing diagrams suggested that molecules

are self-assembled in solid state via non-covalent (C-H···F, C-H···Cl, C-H···O, C-H···S,

C-H···π, F···π and Cl···π interactions to form fascinating supramolecular architectures.

146

Anticancer screening results against different cancer cell lines revealed that most of the

studied complexes have better anticancer activity than the standard drugs doxorubicin

and cisplatin. Like pyriplatin and phenanthriplatin, steric hindrance from C-H groups of

organophosphine plays an important role for the safe entree of these complexes to the

DNA, a critical target for anticancer Pt-drugs. Furthermore, presence of the lipophilic

substituents in the organophosphines enhances the activity by facilitating the cell

internalization process. Similarly, presence of fluoro moiety not only upsurges the

lipophilicity, but also stabilizes the complex-DNA adduct through H-bonding. The results

of DNA-binding studies indicated that the complexes-DNA interaction is mainly non-

intercalative, ensuing by chloride dissociation. A slow chloride exchange with the bulky

ligands (thiourea and diethyldithiocarbamate) than the smaller ones such as thiocyanate,

bromide and iodide is an indication that these complexes are relatively inert to S-

containing off-target biomolecules. The results of DNA cleavage assay showed that the

conversion of super coiled DNA to nicked circular DNA, and the cleavage ability

increases with concentration.

147

LIST OF PUBLICATIONS

1. Amir, M. K.; Zia-ur-Rehman.; Hayat, F.; Khan, S. Z.; Hogarth, G.;

Kondratyuk, T.; Pezzuto, J. M.; Tahir, M. N., Monofunctional platinum (ii)

dithiocarbamate complexes: synthesis, characterization and anticancer

activity. RSC Advances 2016, 6, 110517-110524.

2. Amir, M. K.; Zia-ur-Rehman.; Khan, S. Z.; Hayat, F.; Hassan, A.; Butler, I.

S., Anticancer activity, DNA-binding and DNA-denaturing aptitude of

palladium (II) dithiocarbamates. Inorganica Chimica Acta 2016, 451, 31-40.

3. Khan, S. Z.; Amir, M. K.; Abbasi, R.; Tahir, M. N.; Zia-ur-Rehman, New 3D

and 2D supramolecular heteroleptic palladium (II) dithiocarbamates as potent

anticancer agents. Journal of Coordination Chemistry 2016, 69, 2999-3009.

4. Khan, S. Z.; Amir, M. K.; Ullah, I.; Aamir, A.; Pezzuto, J. M.; Kondratyuk,

T.; Bélanger‐Gariepy, F.; Ali, A.; Khan, S.; Zia-ur-Rehman, New heteroleptic

palladium (II) dithiocarbamates: synthesis, characterization, packing and

anticancer activity against five different cancer cell lines. Applied

Organometallic Chemistry 2016, 6, 392-398.

5. Khan, S. Z.; Amir, M. K.; Naseer, M. M.; Abbasi, R.; Mazhar, K.; Tahir, M.

N.; Awan, I. Z.; Zia-Ur-Rehman, Heteroleptic Pd (II) dithiocarbamates:

synthesis, characterization, packing and in vitro anticancer activity against

HeLa cell line. Journal of Coordination Chemistry 2015, 68, 2539-2551.

6. Amir, M. K.; Zia-ur-Rehman.; Khan, S.; Shah, A.; Butler, I. S., Anticancer

activity of organotin (IV) carboxylates. Inorganica Chimica Acta 2014, 423,

14-25.

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