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Research ArticleBiological Impact of Pd (II) Complexes Synthesis SpectralCharacterization In Vitro Anticancer CT-DNA Binding andAntioxidant Activities

Nitin Kumar Sharma Rakesh Kumar Ameta and Man Singh

School of Chemical Sciences Central University of Gujarat Gandhinagar 382030 India

Correspondence should be addressed to Man Singh mansingh50hotmailcom

Received 12 October 2015 Revised 22 December 2015 Accepted 20 January 2016

Academic Editor Arie Zask

Copyright copy 2016 Nitin Kumar Sharma et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

A new series of Pd (II) complexes of methyl substituted benzylamine ligands (BLs) has been synthesized and characterized viaspectroscopic techniques such as UVVis FTIR LCMS 1H and 13C NMR The UVVis study in DMSO DMSO + water andDMSO + PBS buffer (pH = 72) confirmed their molecular sustainability in liquids Their in vitro anticancer activity against breastcancer cell lines such asMCF-7 andMDA-MB-231 makes them interesting for in vivo analysisTheir stronger DNA binding activity(DBA) compared with free ligand suggested them as a good DNA binder DBA was further confirmed by physicochemical studiessuch as surface tension and viscosity of complex + DNA which inferred the disruption of DNA and intercalation of complexesrespectively Their binding activity disruption of DNA base pairs (DNABP) and intercalating strength are reported inthis paper for the first time for better understanding of DNA binding mechanism Along with this their scavenging activity (SA)determined through DPPH free radical and the results indicate good antioxidant behaviour of complexes

1 Introduction

From the last few decades the transition metal complexeswith different amine ligands have drawn a significant interestin exploring anticancer activities specifically related to solidtumor chemotherapies [1ndash4]The breast cancer a solid tumoris one of the major issues in health care in terms of morbiditymortality and therapy costs [5] To overcome these problemsmany more drugs have been introduced into the marketbut their response to therapy is still very poor Howeversuch drugs have not been found to be much effective fortreatment of solid tumors due to their pertinent side effectssuch as nephrotoxicity drug resistance and renal and cervicalproblems [6 7] Therefore the foremost target of mostresearch groups is to find a convenient anticancer drug thatcan be used efficiently for the treatment of solid tumorsRecently many Pd (II) complexes with promising anticanceractivity against tumor cell lines have been synthesized andreported elsewhere [8] In such studies a good correlationwas observed between the cytostatic activity and lipophilicity

of the Pd complexes [6ndash8] In fact the Pd complexes as anonplatinum complex have recently been reviewed to have asignificant antitumor activity and lower side effects comparedto cisplatin [5] As an essential feature of metal-containinganticancer agents Pd complexes are expected to have lesskidney toxicity than cisplatin [7 8] Further many new Pdcomplexes with amine ligands having promising anticanceractivities with lower side effect have recently been reported[5ndash12] Bearing in mind that Pd (II) complexes are about 105times more reactive than their Pt (II) analogues the lowerantitumor activity of Pd compounds has been attributed tovery rapid hydrolysis of the leaving groups that dissociatereadily in solution leading to reactive species far from theirpharmacological targets [9 10]

Keeping above inconveniences a new series of Pd (II)complexes has been synthesized with methyl substitutedBLs and analyzed their in vitro antitumor activity on breastcancer cell lines MCF-7 and MDA-MB-231 which expressedeffective anticancer potential Since DNA is a primarymolec-ular target of anticancer drugs and ascertains an extent of

Hindawi Publishing CorporationInternational Journal of Medicinal ChemistryVolume 2016 Article ID 9245619 10 pageshttpdxdoiorg10115520169245619

2 International Journal of Medicinal Chemistry

drugrsquos chemotherapeutic potential the DBA of synthesizedcomplexes have chosen to investigate their anticancer nature[8 13] For better understanding of DBA we have alsocalculated the binding activity disruption of DNABPand intercalating strength of complexes by using newlydeveloped quantitative equation Apart from DNA bindingthe antioxidant activity has also proven the anticancer natureand medicinal significance of the Pd (II) complexes whichhave been the criteria for studying their antioxidant activity[14ndash16] Therefore our study of Pd (II) complexes leads toa better understanding of their biological and medicinalapplications

2 Experimental Section

21 Materials and Methods Palladium dichloride (PdCl2)

benzylamine ligands (BLs) CT-DNA Tris buffer DMSO andethanol (gt995) were procured from Sigma-Aldrich andused as received Elemental analysis was made with a Eurovector CHN analyzer andUVVis spectra were recorded witha Spectro 2060 plus spectrophotometer over 200ndash600 nm byusing 1 cm path length cuvette FTIR (Perkin Elmer) spectrawere taken with KBr palate where polystyrene thin film wasused as a calibration standard 1H and 13CNMR spectra wererecorded inDMSO-119889

6(NMR 9999)with a Bruker-Biospin

Avance-III 500MHz FT-NMR spectrometer Mass spectrawere obtained with PE SCIEX API 165 with +ve ESI modewith ammonium acetate and acetonitrile in 1 9 vv ratio asmobile phase Their molecular sustainability was determinedin DMSO DMSOwater and DMSOphosphate buffer ofpH = 72 Buffer solution was prepared by adding 70mL 01MaqueousNaOH into 01M aqueousKH

2PO4solutionThe pH

of a resultant bufferwas checkedwith RS-232modelledCyberscan pH 2100 EUTECH pH meter

22 General Consideration for Synthesis Initially PdCl2and

BLs were separately dissolved in freshly prepared aqueousethanol (absolute ethanol and Milli-Q water in 1 15) in 1 2molar ratio using 1MLH magnetic stirrer The BLs solutionswere added dropwise in metal compound solution with con-tinuous stirring at room temperature After 16 h the mixtureturned from light brown to greenish for Pd complexes andprecipitates were formed The ppts were filtered off andwashed several times with chilled waterethanol in 1 1 ratioand kept overnight in vacuum oven at room temperature forabsolute dryness

23 Characterization Data

231 Complex 1 C16H20N2Pd [Pd2MBA] Yield 01810 g7476 Elemental analysis found C 5574 H 526 N813 Calcd for C

16H20N2Pd C 5582 H 544 N 822

IR (KBr) 120592maxcmminus1 3300 and 32142 (NH

2) 1493 and

14497 (Ph C=C) 7407 (mono substituted Ph) 10519 (CndashN) 4796 (PdndashN) 1H NMR (125MHz DMSO-119889

6 Me4Si)

120575 209 (2H s PhCH2NH2) 393 (2H s PhCH

2NH2) 250

(3H s PhCH3) 737ndash743 (1H d PhH 119869 = 139Hz) 745ndash

747 (1H d PhH 119869 = 73Hz) and 735ndash738 (1H m PhH)13C NMR (125MHz DMSO-119889

6 Me4Si) 120575 4767 (C1) 13652

(C2) 14572 (C3) 12999 (C4) 12883 (C5) 12589 (C6)12740 (C7) and 1875 (C8) +ve ESI-MS mz 3469 [M + 1](calc for [C

16H20N2Pd] = 3747) UVVis in DMSO 120582max [120576

(dm3molminus1cmminus1)] = 275 (2569) 330 (664) 370 (477) nm inDMSO water (1 1) 120582max [120576 (dm

3molminus1cmminus1)] = 270 (2921)340 (358) nm in DMSO phosphate buffer (1 1) 120582max [120576(dm3molminus1cmminus1)] = 265 (2252) 340 (410) nm


16H20N2Pd C 5579 H 533 N 819


2) 1493 and 14497

(Ph C=C) 74464 (mono substituted Ph) 1099 (CndashN) 43742(PdndashN) 1HNMR (500MHz DMSO-119889

6 Me4Si) 120575 2086 (2H

s PhCH2NH2) 3932 (2H s PhCH

2NH2) 2404 (3H s

PhCH3) 721ndash723 (1H d PhH 119869 = 72Hz) 708ndash710 (1H

d PhH 119869 = 72) and 712ndash720 (1H s PhH) 13C NMR(125MHz DMSO-119889

6 Me4Si) 120575 47674 (C1) 13836 (C2)

13723 (C3) 12904 (C4) 12790 (C5) 12548 (C6) 12815 (C7)and 2098 (C8) +ve ESI-MS mz 34605 [M + 1] (calcfor [C




233 Complex 3 C16H20N2Pd [Pd4MBA] Yield 01785 g7420 Elemental analysis found C 5574 H 526 N 813Calcd for C

16H20N2Pd C 5588 H 529 N 821 IR (KBr)

120592maxcmminus1 3257 and 3210 (NH

2) 1453 and 1355 (Ph C=C)

75646 (mono substituted Ph) 10007 and 10361 (CndashN) 4926(PdndashN) 1HNMR (500MHzDMSO-119889

6Me4Si) 120575 209 (2H s

PhCH2NH2) 393 (2H s PhCH

2NH2) 231 (3H s PhCH

3)

733ndash735 (1H d PhH 119869 = 60Hz) 718ndash720 (1H d PhH119869 = 1072Hz) and 725 (1H s PhH) 13C NMR (125MHzDMSO-119889

6 Me4Si) 120575 4767 (C1) 13911 (C2) 12789 (C3) 12876

(C4) 13823 (C5) 12911 (C6) 12814 (C7) and 2029 (C8)+ve ESI-MS mz 3469 [M + 1] (calc for [C

16H20N2Pd] =

34475) UVVis in DMSO 120582max [120576 (dm3molminus1cmminus1)] = 280

(2208) 340 (447) 370 (368) nm in DMSO water (1 1)120582max [120576 (dm3molminus1cmminus1)] = 265 (2770) 340 (286) nm inDMSO phosphate buffer (1 1) 120582max [120576 (dm

3molminus1cmminus1)] =265 (2276) 340 (371) nm

3 Biological Evaluation

31 In Vitro Anticancer Activity Cell viability was esti-mated colorimetrically using 2-(3-diethylamino-6 diethyla-zaniumylidene-xanthen-9-yl)-5-sulfobenzenesulfonate SRB(sulforhodamine B) as standard assay with high reproducibil-ity [17]

311 Cell Lines and Culture Conditions Human breast cancercell lines MCF-7 and MDA-MB-231 were obtained fromNCI USA and grown in minimal essential medium (MEM)Eagles media were supplemented with 10 heat inactivatedfetal bovine serum (FBS Sigma-Aldrich) 2mML-glutamineand 1mm sodium pyruvate (Hyclone) in humidified CO

2

incubator

International Journal of Medicinal Chemistry 3

312 Assay of Cytotoxicity in Cancer Cell Lines Cytotoxicityof Pd (II) complexes was determined by SRB assay whereE5000 cells were seeded into each well of a 96-well clear flatbottom polystyrene tissue culture plate and incubated for 2 hin MEM An additional 190mL cell suspension was added ineach well containing 10mL test sample in 10 DMSO with10mL Adriamycin (doxorubicin) as a positive drug controlEach experiment was carried out in 3 replicate wells After anincubation of 48 h 100mL of 0057 SRB solution (wv) wasadded in each well Then 200mL of 10mM Tris base solution(pH = 105) was added into each well and shaken smoothlyThe cell viability was assayed by absorption at 510 nm with amicroplate readerThe experiments were repeated thrice with3 replicates each time and 99 reproducibility was obtained

32 DNA Binding CT-DNA (analytical grade Sigma-Aldrich) was used as received Tris-HCl buffer (10mM pH= 72) was prepared in Milli-Q water for 50 120583M DNA stocksolution and used for absorption titration viscosity surfacetension conductivity and zeta potential measurements

321 Absorption Spectroscopy DNA concentration wasdetermined using an absorption spectrophotometer thatgave 6600Mminus1cmminus1 a molar absorption coefficient at260 nm [18 19] The CT-DNA in buffer gave a ratio of UVabsorbance at 260 and 280 nm which is found to be 18 to 19and indicates a presence of protein free DNA [20ndash22] The10 30 50 70 and 90 120583M complex solutions were preparedin 10 DMSO in Tris buffer for Pd(BLs)

2to which the DNA

stock solutions were added Their ri = [complex][DNA]at 02 06 1 14 and 18 were calculated by absorptiontitration Before recording UV spectra the Pd(BLs)

2+ DNA

solutions were incubated for 15 min at rt to enable sufficientand reproducible DNA binding with complexes Furthertheir binding strength an intrinsic binding constant (119870

119887)

with CT-DNA was obtained by monitoring a change inDNA absorbance on increasing Pd(BLs)

2concentrations and

calculated with the following equation [23]

[DNA]120576119886minus 120576119887

=[DNA]120576119887minus 120576119891

+

1

119870119887(120576119887minus 120576119891)

(1)

120576119886 120576119891 and 120576

119887are apparent free and bound complex extinc-

tion coefficients respectively 120576119891was determined from a

calibration curve of an isolated metal complex with Beer-Lambert law 120576

119886was determined as a ratio of the measured

absorbance and Pd (II) complexes concentration similarto 119860obs[Pt] A plot of [DNA](120576

119886minus 120576119891) versus [complex]

produced a slope of 1(120576119887minus 120576119891) and a 119884 intercept equal to

1119870119887(120576119887minus 120576119891) 119870119887is the ratio of slope to the 119884 intercept [23]

322 Physicochemical Analysis

(1) Viscosity and Surface Tension Viscosity and surfacetension measurements were made using BMS at 29815 Kand the temperature controlled through an auto temperaturecontrol LAUDA ALPHA RA 8 thermostat [24 25] About15 to 20 measurements were made for each compositionenabling high reproducibility and precision from which

viscous flow time (VFT) and pendent drop numbers (PDN)were calculated (1205781205780)13 versus binding ratio was calculatedwhere 120578 represents the dynamic viscosity of DNA withcomplexes while 1205780 is the viscosity of a DNA mixture inbuffer

(2) Conductance and Zeta Potential Conductance and zetapotentials of DNA solutions with and without complexeswere measured using LABINDIA PICO + conductivityand Microtrac Zetatrac U2771 and DLS respectively at25∘C Aqueous KCl at 01 001 and 0001M of 1288 1413and 147mScmminus1 respectively were used for calibration ofconductivity meter Similarly an auto suspended solution ofalumina suspension (400-206-100) was used as zeta potentialstandard Initially a set-zero was made with DMSO + Trisbuffer for Pd(BLs)

2 The DNA concentration for all measure-

ments was kept constant while the concentration of Pd(BLs)2

was varied from 10 to 90120583M over 20120583M intervals

33 Scavenging Activities Antioxidant activities have beenstudied on free radical scavenging of stable 1-25-diphenyl-2-picrylhydrazyl (DPPH∙) [26] For this purpose stock solutionof complexes and DPPH∙ (0002) were mixed in DMSO +water (1 1) for Pd(BLs)

2 For sample preparation the DPPH∙

solution was mixed with a complex solution in 1 1 ratiofollowed by vigorous shaking and thereafter kept in darkfor 30min incubation The UV absorbance was measuredat 517 nm with UVVis spectrophotometer Later the radicalscavenging activity was measured and a decrease in DPPHabsorbance indicated a radical scavenging activity calculatedwith the following equation

Scavenging activity = (1198600minus 119860119904

1198600

) times 100 (2)

The119860119904is absorbance of DPPH∙ with a test compound and119860

0

absorbance of DPPH∙ without a test compound Absorbancedata are presented as means plusmn SD of three determinations

4 Results and Discussion

41 Synthesis and Characterizations Initially the PdCl2and

BLs ligands were mixed in 1 2 molar ratio for the synthesis ofPd(BLs)

2complexes (Figure 1) The reaction was conducted

in aqueous ethanol solution for 14 to 16 h as per reactionscheme given below

Synthesis of Pd (II) Complexes Consider

PdCl2+ 2BLs

Aqueous C2H5OH

997888997888997888997888997888997888997888997888997888997888997888997888997888rarr

16 hRTPd (BLs)

2 (3)

The 3300 to 3119 cmminus1 stretching frequencies inferred pres-ence of ndashNH

2of benzylamine in the Pd(BLs)

2and similarly

from 1497 to 1453 cmminus1 predicted C=C in phenyl ring The49592 to 43878 cmminus1 and 380ndash348 cmminus1 indicates PdndashNcoordinate and PdndashCl bands respectively [26 27] In 1HNMR the 2Hof ndashNH

2and PhCH

2ndash appeared at 120575 393 to 209

with singlet for Pd(BLs)2 The PhCH

3proton of benzylamine

appeared at 120575 250 to 231 with singlet for all Pd(BLs)2[28]


2-Methylphenylmethanamine 3-Methylphenylmethanamine 4-Methylphenylmethanamine

7

65

4

32

1

8

NH2

7

65

4

32

1

8

NH2

7

65

4

3

2

1

8

NH2

Figure 1 Structures of the ligands (L = BLs)

Two doublets at 120575 737ndash742 and 749ndash747 having 119869 = 139and 73MHz respectively in Pd2MBA for H of C4 and C6appeared Similarly a singlet for 1H of C3 at 120575 720 valueand doublets for 1H of C5 and C6 at 120575 708ndash710 and 721ndash723 (119869 = 72MHz) respectively were found for Pd3MBAA doublet at 120575 733ndash735 and 718ndash720 with 119869 = 60 and1072 MHz respectively was confirmed for 1H of C3 and C4positions while a singlet was confirmed for 1H of C6 at 120575725 in Pd4MBA In 13C NMR the benzyl carbon (PhCH

2ndash)

appears at 120575 4766 for all the Pd(BLs)2[28] The aromatic

ortho meta and para ndashCH3attached carbon appeared at

120575 14572 129036 and 13823 respectively for the Pd(BLs)2

The carbon of ndashCH3at ortho meta and para appeared

within 120575 2098 to 1872 ppm The +ve ESI mass spectra ofPd(BLs)

2have found [M + 1] confirming their molecular

mass In UV study the absorption spectrum consists of aband at 400 nm and may be assigned as 1A

1g rarr 1A2g(119889119909119910rarr 1198891199092minus1199102) transition occurring with Pd complexes (see

ESI Figures 1ndash3 in Supplementary Material available onlineat httpdxdoiorg10115520169245619) [29] To investigatetheir molecular stability in solution the UVVis spectralbehaviour was investigated in DMSO DMSO + water andDMSO + phosphate buffer for Pd(BLs)

2(ESI Figures 1ndash3)

The overall patterns of spectra for complexes solution werefound to be similar to the different mediums to ensure theirstability Thus our synthesized Pd (II) complexes havingUVVis absorption from 265 to 270 nm and 1H NMR cou-pling constant between 5 and 10MHz have confirmed theirtrans geometry (Figure 2)

42 Anticancer Activity Moving towards the higher anti-cancer potential of Pd (II) complexes many of the in vitrostudies on different cell lines have been reported suggestingtheir cytotoxicity up to some extent [30ndash33] In such studiesauthors have synthesized some Pd complexes with differentamine ligands and report their cytotoxicity when tested invitro [30ndash33] It seems that these amine ligands and theirderivatives can further improve the cytotoxicities when theyreacted with different metals Therefore in our study wetried to focus on the factors namely nature of ligands andmethyl group substitution on ligand which can modulatethe cytotoxicity of such complexes In this context we havedemonstrated an in vitro anticancer study of some novel Pdcomplexes using BLs as ligands for better anticancer activityThe complexes were tested in vitro against MCF-7 andMDA-MB-231 human breast tumor cell lines by colorimetricmicroculture 2-(3-diethylamino-6-diethylazaniumylidene-xanthen-9-yl)-5-sulfobenzenesulfonate (SRB) assay and

Pd2MBA Pd3MBA Pd4MBAFigure 2 Structure of synthesized Pd (II) complexes

minus60

minus40

minus20

0

20

40

60

80

100

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBSADR

MCF-7 cell line

Drug concentration (120583M)

Figure 3 Growth curve human breast cancer cell line (MCF-7)

compared with Adriamycin (ADR) and cisplatin [6 34]The MCF-7 and MDA-MB-231 cell lines anticancer activitiesfor 10 20 40 and 80 120583M Pd(BLs)

2have been illustrated

in Figures 3 and 4 The analyzed GI50 TGI and LC

50in

120583M have inferred 50 growth inhibition resultant totalgrowth inhibition and a net loss of 50 cells after treatmentrespectively (Table 1)

The GI50

value less than 10 120583M depicts the anticanceractivity with respect to ADR and cisplatin where the closerGI50

values of Pd2MBA Pd3MBA and Pd4MBA fromstandards make them interesting for testing against othercancerous cell lines

43 DNA Binding Activity

431 Spectrophotometric Method The DBA explains theanticancer nature of the complex or the drug and can beanalyzed by the absorption spectral study In light of this


Table 1 LC50 TGI and GI

50values (120583gmL) against MCF7 and MDA-MB-231 cell lines of complexes an anticancer analysis

Entry ComplexesDrug concentrations (120583M) calculated from graph

MCF7 MDA-MB-231LC50

TGI GI50

LC50

TGI GI50

1 Pd2MBA gt80 794 275 gt80 gt80 6152 Pd3MBA gt80 756 301 gt80 gt80 4463 Pd4MBA gt80 gt80 784 gt80 gt80 gt804 ADR 792 405 lt10 3985 lt10 lt105 Cisplatin gt30 gt30 lt10 gt30 gt30 lt10

the Pd (II) complexes were mixed with CT-DNA at certainconcentration and shown the changes in absorbance Gen-erally the hypochromism effect in UV study reveals all syn-thesized complexes intercalative binding strength attributedto an interaction with DNA bases [35 36] Similarly ahyperchromic effect ascribes an external contact or a par-tial uncoiling of DNA structure exposing more bases ofDNA may be due to electrostatic binding [37 38] Suchobservations have been noticed in the present study wherea significant hypochromic effect in absorption titrations ofDNAwith Pd (II) exposed an intercalationwith the base pairsof DNA [37 38] (ESI Figures 4ndash6)

The decrease in UV absorption predicted the stronginteractions of Pd2MBA Pd3MBA and Pd4MBA whoseintercalating strengths depend on size and electron densitiesof interacting aromatic rings with an amine group [35ndash38]Their DBA has been found stronger in comparison to a freeligand like benzylamine (ESI Figure 7)

Comparative Binding Affinity of Complexes For a betterunderstanding of their binding potential a binding affinity(BA) with DNA has been calculated through well knownspectrophotometric method at a characteristic wavelengthof 260 nm The BA was calculated as per the followingequation (Figure 5)

BA of Complexes = Absorbance of complex at 260 nm minus Absorbance of DNA at 260 nmAbsorbance of DNA at 260 nm

times 100 (4)

Figure 4 shows the BA of each complex where negative signexplains their strong binding with DNA and hypochromismof the complexes

The all Pd (II) complexes show approximately the samebinding activity at each concentration indicating the samebinding potential of each complexes The same bindingpotential can further be explained by the same geometry ofthe all complexesTherefore it can be understood that the Pdcomplexes with the same geometry may equally participatewhen they interacted with DNA

432 Physicochemical Method Along with spectrophoto-metric study the physicochemical study such as viscositysurface tension conductivity and zeta potentials is also aneffective method for better understanding of DBA Thuswe have mathematically established a comparison of theirphysicochemical data that presumes their binding affinitywith DNA at structural level

(1) Study of Structural Disruption of CT-DNA Initially thecomplex entertains the DNA and attacks on its base pairstherefore it weakens the intermolecular forces of baseswhich results in the disruption of base pairs Since it isan intermolecular force disruption and such disruption canbe measured by the analysis of surface tension thereforewe have analysed and compared the surface tensions of

DNA with and without complexes Generally the decreasein surface tension infers the force disruption and we havealso obtained such results where we compared the surfacetension of pureDNAandDNA+ complexes For instance thesurface tension values of the DNA solution with increasingamounts of Pd (II) complexes (1119877 = 02 06 10 14 18) havedecreased as compared to pure DNA (Figure 6)

A decrease in surface tension of Pd (II) complexes +DNAinferred the weakening of intermolecular interaction of DNAbases and has lost interaction with complexes

Complex Contributions on Disruption of DNA Base PairsSurface tension is an important analysing aspect for the dis-ruption of DNA base pairs therefore for the advanced studywe have also measured and compared the disruption ofDNA base pairs The complex contribution was calculatedon the disruption of DNA (Figure 7)

disruption of DNA base pairs

=

120574 of DNA with complexes minus 120574 of pure DNA120574 of pure DNA

times 100

(5)

Figure 7 shows the disruption of DNABP with respect tocomplexes and with negative values indicating the weakening


minus100minus80minus60minus40minus20

020406080

100120

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBAADR

MDA-MB-231 cell line


Figure 4 Growth curve human breast cancer cell line (MDA-MB-231)

minus45minus40minus35minus30minus25minus20minus15minus10minus5

010 30 50 70 90

Bind

ing

activ

ity (

)

Concentration (120583M)

Pd2MBAPd3MBAPd4MBA

Figure 5 binding affinity of complexes at 260 nm

interaction of DNA bases towards disruption This negativevalues support the binding affinity (BA) of complexes wherethe lower ndashve values of Pd complexes indicate the disruptionof DNABP

(2) Study on Intercalation Phenomenon of Complexes Forthe classical intercalative mode the viscosity of the DNAsolution is increased due to an increase in overall DNA lengthwhile it is decreased for a partial nonclassical intercalationprocess causing a bend or kink in the DNA helix that reducesits effective length concomitantly [39 40] For instance adecrease in relative viscosity with cisplatin is explained dueto covalent binding and shortening in an axial length ofdouble helix DNA On the other hand classical organicintercalator such as ethidium bromide increased the relativeviscosity with increasing of an axial length of the DNA[41ndash43] To observe covalent binding or classical organicinteraction their relative specific viscosities (1205781205780)13 (1205780 and120578 being specific viscosity contributions of DNA with andwithout complexes resp) were plotted against 1119877 (119877 =[DNA][complex] = 02 06 10 14 18) (Figure 6)

6100

6132

6164

6196

6228

6260

6292

6324

103

104

106

107

108

110

111

0 03 06 09 12 15 18

Surfa

ce te

nsio

n (m

Mm

)

Pd2MBAPd3MBAPd4MBA

ri = [complex][DNA]

(120578120578

0)13

Figure 6 Effect of increasing amounts of Pd based benzylaminecomplexes on viscosity and surface tension of CT-DNA (5times 10minus5M)

minus1400

minus1325

minus1250

minus1175

minus1100

minus1025

10 30 50 70 90

Disr

uptio

n of

DN

ABP

()

Complex concentration (120583M)

Pd2MBAPd3MBAPd4MBA

Figure 7 disruption of DNA base pairs interaction

On increasing concentration of Pd(BLs)2 an increase

in viscosity of DNA was observed proving the Pd(BLs)2

as DNA intercalators (Figure 6) For ethidium bromide therelative viscosity of DNA is increased with a slope from 0 to09448 LmM [44] whereas with 0001 the relative viscosityof DNA has increased for Pd2MBA Pd3MBA and Pd4MBAThe viscosity of free ligand (2-methylbenzylamine) withDNA was measured and found lower than the complexes +DNA which inferred that interaction of DNA with freeligand is weaker as compared to complexTherefore a higherincrease in viscosity of Pd complex + DNA as comparedto free ligands clearly explained the stronger intercalationbetween complexes and DNA [42] The constant value of theslope and increasing values of viscosity infer almost similarintercalating nature for all the complexes


Complex Contributions towards Intercalation with DNA BasePairs Apart from the above discussion we have also estab-lished thebinding affinitywith respect to viscosity (120578) usingtwodifferentmethods In bothmethods the complex on theintercalation with DNA was calculated

Intercalating strength of complexes

=


times 100

(6)

Figure 8 clearly explains the individual BA of complexeswith DNA base pairs (DNABP) The Pd2MBA Pd3MBAand Pd4MBA expressed lower BA when they interacted withDNA

(3) Conductivity and Zeta Potential The DNA and complexesare charge molecules therefore their conductivity and zetapotential have also been considered as interaction parame-ters as well as key factors for an analyzing conformationalbehaviour of an isolated DNA chain with the complexesA DNA solution of 50 120583M had 3426 120583S conductivity whilewith increasing amounts of complexes the decrease in zetapotential as well as conductivities has been found (Figure 9and Table 2) DNA molecules are negatively charged due tophosphate groups but due to interaction with complexestheir negative charge density is decreased due to Columbicinteraction of positively charged complexes and the disasso-ciation of counter ions in DNA

Thedecrease in conductivity and zeta potential confirmedthat the DNA could structurally be modified due to suchinteractions

44 Drug Efficacy Studies or DFI with DNA Binding For thebetter understanding of Pd(BLs)

2+ DNA interaction and

their disruption during an intercalation process DFI hasfound reliable data in support of anticancer activity [42]Therefore our study infers that the Pd(BLs)

2+ DNA inter-

actions at different concentration have shown their higherviscosities and lower surface tension with respect to DNAsolution (Figure 6) Thus a disruption of DNABP decreasessurface tension by developing an interaction between com-plexes and DNA

45 Scavenging Activities The scavenging activities havebeen investigated to support the anticancer potential of suchcomplexes [43 44] Recently it is reported that by usinghydroxyl radicals an inorganic radical (most dangerousreactive oxygen species) with its precursor such as H

2O2

proposed the facts responsible for their mode of mechanism[43] On the basis of the above study the antioxidant activitieshave studied and analyzed the decrease in absorbance orscavenging effect of a stable free DPPH∙ as per standard pro-cedure for the Pd(BLs)

2[45 46] The percentage scavenging

activity of Pd(BLs)2has been determined in a concentration-

dependent mode in comparison with the DPPH∙ absorptionat 517 nm [47ndash50] The DPPH∙ free radicalrsquos absorption at517 nm with DMSO was 0906 and for Pd(BLs)

2 From 50 to

Table 2 Conductivity (120583S) of DNA and DNA + complexes

Conductivity120583S000lowast 10 30 50 70 90

Pd2MBA 3426 3053 3013 2974 2903 2898Pd3MBA 3426 2988 2978 2966 2952 2912Pd3MBA 3426 2761 2687 2645 2581 2555lowast refers to 50120583MDNA

700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA


Figure 8 intercalating strength of complexes with DNA basepairs

125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA


Figure 9 Zeta potential (mV) of pure DNA and DNA + complexes

250 120583Mwith an interval of 50 120583M complexes have expresseda decrease in absorption (Figure 10) that characterised themas an antioxidant [51]

The 5547 28812 and 2424 for Pd2MBA Pd3MBAand Pd4MBA respectively inferred Pd2MBA a strongantioxidant among them Thus the antioxidant activities ofPd complexes have inferred their significance in medicinalsciences


2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250


Figure 10 Free radical scavenging activities of synthesized com-plexes

5 Conclusion

Since the synthesized Pd(BLs)2could not express the anti-

cancer activity their DBA and SA refer them as anticancercompounds having somemodificationThe closerGI

50values

as compared to control drugs such as doxorubicin andcisplatin make the synthesized complexes more interesting totest against other cancerous cell lines The DBA analysis withviscosity surface tension zeta potential and conductance hasinferred strong intercalating as well as DNA disruption andintercalating nature of the complexes The quantitative eval-uation of DNABP disruption and intercalating strengthadvances the DNA binding mechanism The complexes haveshown significant free radical scavenging activities actingas antioxidants and could be used for medicinal purposesThereby our study could have potential in better understand-ing of medicinal aspects of the complexes

Supporting Information

It includes UVVis spectra of stability DNA binding andconductivities (DNA + complexes) of the complexes

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

The authors are thankful to Central University of GujaratGandhinagar for financial infrastructural support andexperimental facilities

References

[1] P J Loehrer and L H Einhorn ldquoDrugs five years laterCisplatinrdquo Annals of Internal Medicine vol 100 no 5 pp 704ndash713 1984

[2] C A Puckett and J K Barton ldquoMechanism of cellular uptakeof a ruthenium polypyridyl complexrdquo Biochemistry vol 47 no45 pp 11711ndash11716 2008

[3] K L Haas and K J Franz ldquoApplication of metal coordinationchemistry to explore and manipulate cell biologyrdquo ChemicalReviews vol 109 no 10 pp 4921ndash4960 2009

[4] F Liu W Zhang S He L Wang and X Liu ldquoSynthesiscrystal structure and properties of the binuclear complex[Sm2(C10H9N2O4)2(C10H8N2O4)2(H2O)4]rdquo Journal of Chemi-

cal Crystallography vol 38 no 10 pp 755ndash760 2008[5] J R Benson I Jatoi M Keisch F J Esteva A Makris and V C

Jordan ldquoEarly breast cancerrdquoThe Lancet vol 373 no 9673 pp1463ndash1479 2009

[6] M Galanski ldquoRecent developments in the field of anticancerplatinum complexesrdquo Recent Patents on Anti-Cancer DrugDiscovery vol 1 no 2 pp 285ndash295 2006

[7] E Gao C Liu M Zhu H Lin Q Wu and L Liu ldquoCur-rent development of Pd(II) complexes as potential antitumoragentsrdquo Anti-Cancer Agents in Medicinal Chemistry vol 9 no3 pp 356ndash368 2009

[8] A S Abu-Surrah K A Abu Safieh I M Ahmad et al ldquoNewpalladium(II) complexes bearing pyrazole-based Schiff baseligands synthesis characterization and cytotoxicityrdquo EuropeanJournal of Medicinal Chemistry vol 45 no 2 pp 471ndash475 2010

[9] D S Gill ldquoStructure activity relationship of antitumor pal-ladium complexesrdquo in Platinum Coordination Complexes inCancer Chemotherapy M P Hacker E B Douple and I HKrakoff Eds vol 17 of Developments in Oncology pp 267ndash278Springer New York NY USA 1984

[10] N A Al-Masoudi B H Abdullah A H Essa R Loddo and PLaColla ldquoPlatinum and palladium-triazole complexes as highlypotential antitumor agentsrdquo Archiv der Pharmazie vol 343 no4 pp 222ndash227 2010

[11] A Divsalar A A Saboury H Mansoori-Torshizi and FAhmad ldquoDesign synthesis and biological evaluation of a newpalladium(II) complex 120573-lactoglobulin and K562 as targetsrdquoJournal of Physical Chemistry B vol 114 no 10 pp 3639ndash36472010

[12] G Tamasi M Casolaro A Magnani et al ldquoNew platinum-oxicam complexes as anti-cancer drugs Synthesis character-ization release studies from smart hydrogels evaluation ofreactivity with selected proteins and cytotoxic activity in vitrordquoJournal of Inorganic Biochemistry vol 104 no 8 pp 799ndash8142010

[13] V Mahalingam N Chitrapriya F R Fronczek and K Natara-jan ldquoDimethyl sulfoxide ruthenium(II) complexes of thiosemi-carbazones and semicarbazone synthesis characterization andbiological studiesrdquo Polyhedron vol 27 no 13 pp 2743ndash27502008

[14] J Cao X Xia X Chen J Xiao and QWang ldquoCharacterizationof flavonoids from Dryopteris erythrosora and evaluation oftheir antioxidant anticancer and acetylcholinesterase inhibi-tion activitiesrdquo Food and Chemical Toxicology vol 51 no 1 pp242ndash250 2013

[15] A Huber PThongphasuk G Erben et al ldquoSignificantly greaterantioxidant anticancer activities of 23-dehydrosilybin thansilybinrdquoBiochimica et Biophysica Acta (BBA)mdashGeneral Subjectsvol 1780 no 5 pp 837ndash847 2008

[16] S M M Shanab S S M Mostafa E A Shalaby and G IMahmoud ldquoAqueous extracts of microalgae exhibit antioxidantand anticancer activitiesrdquo Asian Pacific Journal of TropicalBiomedicine vol 2 no 8 pp 608ndash615 2012


[17] P Skehan R Storeng D Scudiero et al ldquoNew colorimetriccytotoxicity assay for anticancer-drug screeningrdquo Journal of theNational Cancer Institute vol 82 no 13 pp 1107ndash1112 1990

[18] P Zhao L-C Xu J-W Huang et al ldquoTricationic pyridiumporphyrins appending different peripheral substituents exper-imental and DFT studies on their interactions with DNArdquoBiophysical Chemistry vol 135 no 1ndash3 pp 102ndash109 2008

[19] P Zhao L-C Xu J-W Huang et al ldquoExperimental and DFTstudies on DNA binding and photocleavage of two cationicporphyrins effects of the introduction of a carboxyphenyl intopyridinium porphyrinrdquo Spectrochimica Acta Part A Molecularand Biomolecular Spectroscopy vol 71 no 4 pp 1216ndash12232008

[20] P Zhao L-C Xu J-W Huang et al ldquoDNA-binding andphotocleavage properties of cationic porphyrinndashanthraquinonehybrids with different lengths of linksrdquo Bioorganic Chemistryvol 36 no 6 pp 278ndash287 2008

[21] J Z Lu Y F Du H W Guo J Jiang X D Zeng andL Q Zang ldquoTwo oxovanadium complexes incorporatingthiosemicarbazones synthesis characterization and DNA-binding studiesrdquo Journal of Coordination Chemistry vol 64 no7 pp 1229ndash1239 2011

[22] Y-F Du J-Z Lu H-W Guo et al ldquoDNA binding andphotocleavage properties of two mixed-ligand oxovanadiumcomplexesrdquo Transition Metal Chemistry vol 35 no 7 pp 859ndash864 2010

[23] N Shahabadi S Kashanian and M Purfoulad ldquoDNA interac-tion studies of a platinum(II) complex PtCl

2(NN) (NN=47-

dimethyl-110-phenanthroline) using different instrumentalmethodsrdquo Spectrochimica Acta Part A Molecular and Biomolec-ular Spectroscopy vol 72 no 4 pp 757ndash761 2009

[24] M Singh ldquoSurvismetermdashtype I and II for surface tension vis-cosity measurements of liquids for academic and research anddevelopment studiesrdquo Journal of Biochemical and BiophysicalMethods vol 67 no 2-3 pp 151ndash161 2006

[25] M Singh ldquoSurvismeter 2-in-1 for viscosity and surface tensionmeasurement an excellent invention for industrial proliferationof surface forces in liquidsrdquo Surface Review and Letters vol 14no 5 pp 973ndash983 2007

[26] Y Sun S Gou R Yin and P Jiang ldquoSynthesisantiproliferative activity and DNA binding study of mixedamminecyclohexylamine platinum(II) complexes with 1-(substituted benzyl) azetidine-3 3-dicarboxylatesrdquo EuropeanJournal of Medicinal Chemistry vol 46 no 10 pp 5146ndash51532011

[27] A D Allen and C V Senoff ldquoPreparation and infraredspectra of some ammine complexes of ruthenium(II) andruthenium(III)rdquo Canadian Journal of Chemistry vol 45 no 12pp 1337ndash1341 1967

[28] N D Ball J W Kampf and M S Sanford ldquoSynthesis and reac-tivity of palladium(II) fluoride complexes containing nitrogen-donor ligandsrdquo Dalton Transactions vol 39 no 2 pp 632ndash6402010

[29] L Trynda D Kwiatkowska andWTyran ldquoPlatinum complexesand pyruvate kinase activityrdquoGeneral Physiology and Biophysicsvol 17 no 1 pp 25ndash36 1998

[30] C Gao F Fei TWang et al ldquoSynthesis and in vitro cytotoxicityof platinum(II) complexes with chiral N-monosubstituted 12-cyclohexyldiamine derivatives as the carrier groupsrdquo Journal ofCoordination Chemistry vol 66 no 6 pp 1068ndash1076 2013

[31] L Li J Zhang L Ma et al ldquoSynthesis characterization andcytotoxicity of platinum(II)palladium(II) complexes with 4-toluenesulfonyl-L-amino acid dianion and diiminediaminerdquoJournal of Coordination Chemistry vol 66 no 4 pp 638ndash6492013

[32] M N Patel P A Dosi and B S Bhatt ldquoSquare planar palla-dium(II) complexes of bipyridines synthesis characterizationand biological studiesrdquo Journal of Coordination Chemistry vol65 no 21 pp 3833ndash3844 2012

[33] J Zhang F Zhang LWang J Du SWang and S Li ldquoSynthesischaracterization and cytotoxicity of complexes of platinum(II)with 221015840-bipyridine and N-benzoyl-L-amino acid dianionrdquoJournal of Coordination Chemistry vol 65 no 12 pp 2159ndash21692012

[34] M Navarro W Castro A R Higuera-Padilla et al ldquoSynthesischaracterization and biological activity of trans-platinum(II)complexes with chloroquinerdquo Journal of Inorganic Biochemistryvol 105 no 12 pp 1684ndash1691 2011

[35] B-D Wang Z-Y Yang P Crewdson and D-Q WangldquoSynthesis crystal structure and DNA-binding studies ofthe Ln(III) complex with 6-hydroxychromone-3-carbaldehydebenzoyl hydrazonerdquo Journal of Inorganic Biochemistry vol 101no 10 pp 1492ndash1504 2007

[36] J K Barton J J Dennenberg and J B ChairesldquoTris(phenanthroline)ruthenium(II) enantiomer interactionswith DNA mode and specificity of bindingrdquo Biochemistry vol32 no 10 pp 2573ndash2584 1993

[37] R F Pasternack E J Gibbs and J J Villafranca ldquoInteractionsof porphyrins with nucleic acidsrdquo Biochemistry vol 22 no 10pp 2406ndash2414 1983

[38] G Pratviel J Bernadou and B Meunier ldquoDNA and RNAcleavage by metal complexesrdquo Advances in Inorganic Chemistryvol 45 pp 251ndash312 1998

[39] J Lu H Guo X Zeng et al ldquoSynthesis and characterizationof unsymmetrical oxidovanadium complexes DNA-bindingcleavage studies and antitumor activitiesrdquo Journal of InorganicBiochemistry vol 112 pp 39ndash48 2012

[40] R K Ameta and M Singh ldquoSAR and DFI studies ofsupramolecular tetraammoniumplatinate + DNA matrix withUVVis spectrophotometry and physicochemical analysis at29815 Krdquo Journal of Molecular Liquids vol 190 pp 200ndash2072014

[41] L Kapicak and E J Gabbay ldquoTopography of nucleic acid helixesin solutions XXXIII Effect of aromatic cations on the tertiarystructures of deoxyribonucleic acidrdquo Journal of the AmericanChemical Society vol 97 pp 403ndash408 1975

[42] E C Long and J K Barton ldquoOn demonstrating DNA intercala-tionrdquo Accounts of Chemical Research vol 23 no 9 pp 271ndash2731990

[43] DMeyerstein ldquoComment on the section lsquoantioxidantmeasure-ments and hydroxyl radical scavenging activityrsquo in synthesischaracterization DNA binding and antioxidant activities offour copper(II) complexes containing N-(3-hydroxybenzyl)-amino amide ligandsrdquo Journal of Coordination Chemistry vol66 pp 2076ndash2078 2013

[44] K Jomova S Baros and M Valko ldquoRedox active metal-induced oxidative stress in biological systemsrdquo TransitionMetalChemistry vol 37 no 2 pp 127ndash134 2012

[45] Z A Taha AM AjlouniW AlMomani and A A Al-GhzawildquoSyntheses characterization biological activities and photo-physical properties of lanthanides complexeswith a tetradentate


Schiff base ligandrdquo Spectrochimica Acta Part A Molecular andBiomolecular Spectroscopy vol 81 no 1 pp 570ndash577 2011

[46] R Trivedi S B Deepthi L Giribabu et al ldquoSynthesis crystalstructure electronic spectroscopy electrochemistry and biolog-ical studies of FerrocenendashCarbohydrate conjugatesrdquo EuropeanJournal of Inorganic Chemistry no 13 pp 2267ndash2277 2012

[47] D Suh and J B Chaires ldquoCriteria for the mode of binding ofDNA binding agentsrdquo Bioorganic and Medicinal Chemistry vol3 no 6 pp 723ndash728 1995

[48] F-H Li G-H Zhao H-X Wu et al ldquoSynthesis character-ization and biological activity of lanthanum(III) complexescontaining 2-methylene-110-phenanthroline units bridged byaliphatic diaminesrdquo Journal of Inorganic Biochemistry vol 100no 1 pp 36ndash43 2006

[49] M E Howe-Grant and S J Lippard ldquoAqueous platinum (II)chemistry binding to biological moleculesrdquo Metal Ions inBiological Systems vol 20 pp 63ndash125 1980

[50] R K AmetaM Singh and R K Kale ldquoSynthesis and structure-activity relationship of benzylamine supported platinum(IV)complexesrdquo New Journal of Chemistry vol 37 no 5 pp 1501ndash1508 2013

[51] A M J Fichtinger-Schepman J L Van der Veer J H J DenHartog P H M Lohman and J Reedijk ldquoAdducts of theantitumor drug cis-diamminedichloroplatinum(II) with DNAformation identification and quantitationrdquo Biochemistry vol24 no 3 pp 707ndash713 1985

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Inorganic ChemistryInternational Journal of

Hindawi Publishing Corporation httpwwwhindawicom Volume 2014

International Journal ofPhotoenergy


Carbohydrate Chemistry

International Journal of


Journal of

Chemistry


Advances in

Physical Chemistry

Hindawi Publishing Corporationhttpwwwhindawicom

Analytical Methods in Chemistry

Journal of

Volume 2014

Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

SpectroscopyInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Medicinal ChemistryInternational Journal of


Chromatography Research International


Applied ChemistryJournal of



Theoretical ChemistryJournal of


Journal of

Spectroscopy

Analytical ChemistryInternational Journal of


Journal of


Quantum Chemistry


Organic Chemistry International

ElectrochemistryInternational Journal of



CatalystsJournal of


drugrsquos chemotherapeutic potential the DBA of synthesizedcomplexes have chosen to investigate their anticancer nature[8 13] For better understanding of DBA we have alsocalculated the binding activity disruption of DNABPand intercalating strength of complexes by using newlydeveloped quantitative equation Apart from DNA bindingthe antioxidant activity has also proven the anticancer natureand medicinal significance of the Pd (II) complexes whichhave been the criteria for studying their antioxidant activity[14ndash16] Therefore our study of Pd (II) complexes leads toa better understanding of their biological and medicinalapplications

2 Experimental Section

21 Materials and Methods Palladium dichloride (PdCl2)

benzylamine ligands (BLs) CT-DNA Tris buffer DMSO andethanol (gt995) were procured from Sigma-Aldrich andused as received Elemental analysis was made with a Eurovector CHN analyzer andUVVis spectra were recorded witha Spectro 2060 plus spectrophotometer over 200ndash600 nm byusing 1 cm path length cuvette FTIR (Perkin Elmer) spectrawere taken with KBr palate where polystyrene thin film wasused as a calibration standard 1H and 13CNMR spectra wererecorded inDMSO-119889

6(NMR 9999)with a Bruker-Biospin

Avance-III 500MHz FT-NMR spectrometer Mass spectrawere obtained with PE SCIEX API 165 with +ve ESI modewith ammonium acetate and acetonitrile in 1 9 vv ratio asmobile phase Their molecular sustainability was determinedin DMSO DMSOwater and DMSOphosphate buffer ofpH = 72 Buffer solution was prepared by adding 70mL 01MaqueousNaOH into 01M aqueousKH

2PO4solutionThe pH

of a resultant bufferwas checkedwith RS-232modelledCyberscan pH 2100 EUTECH pH meter

22 General Consideration for Synthesis Initially PdCl2and

BLs were separately dissolved in freshly prepared aqueousethanol (absolute ethanol and Milli-Q water in 1 15) in 1 2molar ratio using 1MLH magnetic stirrer The BLs solutionswere added dropwise in metal compound solution with con-tinuous stirring at room temperature After 16 h the mixtureturned from light brown to greenish for Pd complexes andprecipitates were formed The ppts were filtered off andwashed several times with chilled waterethanol in 1 1 ratioand kept overnight in vacuum oven at room temperature forabsolute dryness

23 Characterization Data


16H20N2Pd C 5582 H 544 N 822


2) 1493 and

14497 (Ph C=C) 7407 (mono substituted Ph) 10519 (CndashN) 4796 (PdndashN) 1H NMR (125MHz DMSO-119889

6 Me4Si)

120575 209 (2H s PhCH2NH2) 393 (2H s PhCH

2NH2) 250

(3H s PhCH3) 737ndash743 (1H d PhH 119869 = 139Hz) 745ndash

747 (1H d PhH 119869 = 73Hz) and 735ndash738 (1H m PhH)13C NMR (125MHz DMSO-119889

6 Me4Si) 120575 4767 (C1) 13652

(C2) 14572 (C3) 12999 (C4) 12883 (C5) 12589 (C6)12740 (C7) and 1875 (C8) +ve ESI-MS mz 3469 [M + 1](calc for [C





16H20N2Pd C 5579 H 533 N 819


2) 1493 and 14497

(Ph C=C) 74464 (mono substituted Ph) 1099 (CndashN) 43742(PdndashN) 1HNMR (500MHz DMSO-119889

6 Me4Si) 120575 2086 (2H

s PhCH2NH2) 3932 (2H s PhCH

2NH2) 2404 (3H s

PhCH3) 721ndash723 (1H d PhH 119869 = 72Hz) 708ndash710 (1H

d PhH 119869 = 72) and 712ndash720 (1H s PhH) 13C NMR(125MHz DMSO-119889

6 Me4Si) 120575 47674 (C1) 13836 (C2)

13723 (C3) 12904 (C4) 12790 (C5) 12548 (C6) 12815 (C7)and 2098 (C8) +ve ESI-MS mz 34605 [M + 1] (calcfor [C




233 Complex 3 C16H20N2Pd [Pd4MBA] Yield 01785 g7420 Elemental analysis found C 5574 H 526 N 813Calcd for C

16H20N2Pd C 5588 H 529 N 821 IR (KBr)

120592maxcmminus1 3257 and 3210 (NH

2) 1453 and 1355 (Ph C=C)

75646 (mono substituted Ph) 10007 and 10361 (CndashN) 4926(PdndashN) 1HNMR (500MHzDMSO-119889

6Me4Si) 120575 209 (2H s

PhCH2NH2) 393 (2H s PhCH

2NH2) 231 (3H s PhCH

3)

733ndash735 (1H d PhH 119869 = 60Hz) 718ndash720 (1H d PhH119869 = 1072Hz) and 725 (1H s PhH) 13C NMR (125MHzDMSO-119889

6 Me4Si) 120575 4767 (C1) 13911 (C2) 12789 (C3) 12876

(C4) 13823 (C5) 12911 (C6) 12814 (C7) and 2029 (C8)+ve ESI-MS mz 3469 [M + 1] (calc for [C

16H20N2Pd] =

34475) UVVis in DMSO 120582max [120576 (dm3molminus1cmminus1)] = 280

(2208) 340 (447) 370 (368) nm in DMSO water (1 1)120582max [120576 (dm3molminus1cmminus1)] = 265 (2770) 340 (286) nm inDMSO phosphate buffer (1 1) 120582max [120576 (dm

3molminus1cmminus1)] =265 (2276) 340 (371) nm

3 Biological Evaluation

31 In Vitro Anticancer Activity Cell viability was esti-mated colorimetrically using 2-(3-diethylamino-6 diethyla-zaniumylidene-xanthen-9-yl)-5-sulfobenzenesulfonate SRB(sulforhodamine B) as standard assay with high reproducibil-ity [17]

311 Cell Lines and Culture Conditions Human breast cancercell lines MCF-7 and MDA-MB-231 were obtained fromNCI USA and grown in minimal essential medium (MEM)Eagles media were supplemented with 10 heat inactivatedfetal bovine serum (FBS Sigma-Aldrich) 2mML-glutamineand 1mm sodium pyruvate (Hyclone) in humidified CO

2

incubator





2to which the DNA


2+ DNA


119887)


2concentrations and


[DNA]120576119886minus 120576119887

=[DNA]120576119887minus 120576119891

+

1

119870119887(120576119887minus 120576119891)

(1)

120576119886 120576119891 and 120576




















1198600

) times 100 (2)


0








PdCl2+ 2BLs

Aqueous C2H5OH

997888997888997888997888997888997888997888997888997888997888997888997888997888rarr

16 hRTPd (BLs)

2 (3)



2and similarly


2and PhCH







7

65

4

32

1

8

NH2

7

65

4

32

1

8

NH2

7

65

4

3

2

1

8

NH2



2ndash)














minus60

minus40

minus20

0

20

40

60

80

100

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBSADR

MCF-7 cell line






50in


The GI50









MCF7 MDA-MB-231LC50

TGI GI50

LC50

TGI GI50






times 100 (4)









=


times 100

(5)




020406080

100120

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBAADR





010 30 50 70 90

Bind

ing

activ

ity (

)


Pd2MBAPd3MBAPd4MBA




6100

6132

6164

6196

6228

6260

6292

6324

103

104

106

107

108

110

111

0 03 06 09 12 15 18

Surfa

ce te

nsio

n (m

Mm

)

Pd2MBAPd3MBAPd4MBA

ri = [complex][DNA]

(120578120578

0)13


minus1400

minus1325

minus1250

minus1175

minus1100

minus1025

10 30 50 70 90

Disr

uptio

n of

DN

ABP

()


Pd2MBAPd3MBAPd4MBA








=


times 100

(6)







2+ DNA inter-



2O2





2 From 50 to




700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA



125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA






2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of





2to which the DNA


2+ DNA


119887)


2concentrations and


[DNA]120576119886minus 120576119887

=[DNA]120576119887minus 120576119891

+

1

119870119887(120576119887minus 120576119891)

(1)

120576119886 120576119891 and 120576




















1198600

) times 100 (2)


0








PdCl2+ 2BLs

Aqueous C2H5OH

997888997888997888997888997888997888997888997888997888997888997888997888997888rarr

16 hRTPd (BLs)

2 (3)



2and similarly


2and PhCH







7

65

4

32

1

8

NH2

7

65

4

32

1

8

NH2

7

65

4

3

2

1

8

NH2



2ndash)














minus60

minus40

minus20

0

20

40

60

80

100

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBSADR

MCF-7 cell line






50in


The GI50









MCF7 MDA-MB-231LC50

TGI GI50

LC50

TGI GI50






times 100 (4)









=


times 100

(5)




020406080

100120

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBAADR





010 30 50 70 90

Bind

ing

activ

ity (

)


Pd2MBAPd3MBAPd4MBA




6100

6132

6164

6196

6228

6260

6292

6324

103

104

106

107

108

110

111

0 03 06 09 12 15 18

Surfa

ce te

nsio

n (m

Mm

)

Pd2MBAPd3MBAPd4MBA

ri = [complex][DNA]

(120578120578

0)13


minus1400

minus1325

minus1250

minus1175

minus1100

minus1025

10 30 50 70 90

Disr

uptio

n of

DN

ABP

()


Pd2MBAPd3MBAPd4MBA








=


times 100

(6)







2+ DNA inter-



2O2





2 From 50 to




700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA



125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA






2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of



7

65

4

32

1

8

NH2

7

65

4

32

1

8

NH2

7

65

4

3

2

1

8

NH2



2ndash)














minus60

minus40

minus20

0

20

40

60

80

100

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBSADR

MCF-7 cell line






50in


The GI50









MCF7 MDA-MB-231LC50

TGI GI50

LC50

TGI GI50






times 100 (4)









=


times 100

(5)




020406080

100120

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBAADR





010 30 50 70 90

Bind

ing

activ

ity (

)


Pd2MBAPd3MBAPd4MBA




6100

6132

6164

6196

6228

6260

6292

6324

103

104

106

107

108

110

111

0 03 06 09 12 15 18

Surfa

ce te

nsio

n (m

Mm

)

Pd2MBAPd3MBAPd4MBA

ri = [complex][DNA]

(120578120578

0)13


minus1400

minus1325

minus1250

minus1175

minus1100

minus1025

10 30 50 70 90

Disr

uptio

n of

DN

ABP

()


Pd2MBAPd3MBAPd4MBA








=


times 100

(6)







2+ DNA inter-



2O2





2 From 50 to




700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA



125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA






2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of





MCF7 MDA-MB-231LC50

TGI GI50

LC50

TGI GI50






times 100 (4)









=


times 100

(5)




020406080

100120

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBAADR





010 30 50 70 90

Bind

ing

activ

ity (

)


Pd2MBAPd3MBAPd4MBA




6100

6132

6164

6196

6228

6260

6292

6324

103

104

106

107

108

110

111

0 03 06 09 12 15 18

Surfa

ce te

nsio

n (m

Mm

)

Pd2MBAPd3MBAPd4MBA

ri = [complex][DNA]

(120578120578

0)13


minus1400

minus1325

minus1250

minus1175

minus1100

minus1025

10 30 50 70 90

Disr

uptio

n of

DN

ABP

()


Pd2MBAPd3MBAPd4MBA








=


times 100

(6)







2+ DNA inter-



2O2





2 From 50 to




700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA



125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA






2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of



020406080

100120

10 20 40 80

Con

trol g

row

th (

)

Pd2MBAPd3MBA

Pd4MBAADR





010 30 50 70 90

Bind

ing

activ

ity (

)


Pd2MBAPd3MBAPd4MBA




6100

6132

6164

6196

6228

6260

6292

6324

103

104

106

107

108

110

111

0 03 06 09 12 15 18

Surfa

ce te

nsio

n (m

Mm

)

Pd2MBAPd3MBAPd4MBA

ri = [complex][DNA]

(120578120578

0)13


minus1400

minus1325

minus1250

minus1175

minus1100

minus1025

10 30 50 70 90

Disr

uptio

n of

DN

ABP

()


Pd2MBAPd3MBAPd4MBA








=


times 100

(6)







2+ DNA inter-



2O2





2 From 50 to




700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA



125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA






2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of




=


times 100

(6)







2+ DNA inter-



2O2





2 From 50 to




700

875

1050

1225

1400

1575

1750

10 30 50 70 90In

terc

alat

ing

stren

gth

()

Pd2MBAPd3MBAPd4MBA



125

140

155

170

185

200

DNA 10 30 50 70 90

Zeta

pot

entia

l (m

V)

Pd2MBAPd3MBAPd4MBA






2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


2513

3623

4733

5843

Scav

engi

ng ac

tivity

()

Pd2MBAPd3MBAPd4MBA

50 100 150 200 250



5 Conclusion



50values






Acknowledgment


References


























2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of









2(NN) (NN=47-









































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


















Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of

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