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Hindawi Publishing CorporationISRN Inorganic ChemistryVolume 2013 Article ID 250791 11 pageshttpdxdoiorg1011552013250791

Research ArticleSynthesis Crystal Structure Antioxidant Antimicrobial andMutagenic Activities and DNA Interaction Studies ofNi(II) Schiff Base 4-Methoxy-3-benzyloxybenzaldehydeThiosemicarbazide Complexes

P R Chetana1 M N Somashekar1 B S Srinatha1 R S Policegoudra2

S M Aradhya3 and Ramakrishna Rao4

1 Department of Chemistry Central College Campus Bangalore University Bangalore 560001 India2Department of Pharmaceutical Technology Defence Research Laboratory Tezpur 784001 India3 Department of Fruit and Vegetable Technology CFTRI Mysore 570020 India4Department of Inorganic and Physical Chemistry Indian Institute of Science Sir CV Raman Avenue Bangalore 560012 India

Correspondence should be addressed to P R Chetana prchetanagmailcom

Received 31 July 2013 Accepted 18 August 2013

Academic Editors P Deplano and K Dhara

Copyright copy 2013 P R Chetana et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Three new Ni(II) square planar complexes of 4-methoxy-3-benzyloxybenzaldehyde thiosemicarbazide(4m3BTSC) havingpolypyridyl bases of general formulation [ML

2] (1) and [MLB] (2 3) where L = 4m3BTSC and B is NN-donor heterocyclic bases

namely 110-phenanthroline (phen 2) 221015840-bipyridine (bpy 3) are synthesized and characterizedThe free radical scavenging assayresults showed that complex 1 possesses significant activity when compared to complexes 2 and 3 The biological studies showedthat the ligand and its complexes exhibited significant and different biological activities and also the prepared compounds arenonmutagenic They may be potential commercial antioxidants because of their nonmutagenic and nontoxic nature The DNAinteraction of the new complexes is evaluated by absorption emission andmelting temperature methods and the results suggestedthat the binding affinity of the complexes increases with the presence of planar ligand in the molecule The nickel (II) complexeswith planar phenanthroline bases show moderate DNA binding and cleavage ability

1 Introduction

Schiff bases have great importance in coordination chemistrydue to their ability to form a range of complexes withapplications in different fields [1] Thiosemicarbazones havevariable bonding modes ability to form stable chelates withmetal ions and structural diversity [2] The sulfur oxygenand nitrogen may be involved in coordination providing auseful model for bioinorganic processes [3] These thiosemi-carbazide ligands have good complexing ability and theiractivity increases on complexation with transition metal ions[4] These compounds are also of interest due to their biolog-ical activities including enzyme inhibition [5] and antifungaland pharmacological applications [6 7] The toxicologicalimportance of the ndashNndashC=S moiety has been well established

in antifungal antibacterial agents and pesticidal activities[8 9] It has been suggested that the azomethine linkage inSchiff bases is responsible for the biological activities such asantitumor antibacterial antifungal and herbicidal activities[10]

The biological activities of thiosemicarbazones oftenshowed a high dependence on their substituents Minormodifications in the structure of thiosemicarbazones canlead to widely different biological activities The biologicalproperties of thiosemicarbazones are often modulated bymetal coordination In some cases the highest biologicalactivity is associated with a metal and some side effects maydecrease upon complexation [11]

Nickel(II) complexes have found several potential appli-cations in medicine and very recently they have been

2 ISRN Inorganic Chemistry

screened for inhibition of cancer cell proliferation [12 13]In the last 20 years transition metal complexes have becomeincreasingly important as drugs and artificial nucleasesOne goal of designing Ni(II) complexes is developing morereactive chemical systems that can efficiently cleave DNAunder physiological conditions [14ndash16]

Substantial progress has been made during the past fewdecades to develop metal-based small molecules as DNAfoot-printing as well as the therapeutic agents that are capableof binding and cleaving DNA under physiological condition[17 18] Among the metal complexes so far investigatedthose of polypyridyl phenanthroline bases have attractedgreat attention by virtue of its binding propensity to nucleicacid under the physiological condition [19 20]

In view of the varying ligation behavior and interestingbiological activity shown by these nitrogen-sulfur ligandsand their complexes we are also interested to preparethe thiosemicarbazide Schiff base and its metal complexesDesigning of these metal complexes has been undertakenand their biological activities have been studiedThe study ofbiological activities of benzyloxy benzaldehyde-Schiff basesis less explored Herein we present the syntheses struc-ture antioxidant antimicrobial and mutagenic activitiesand DNA interactions of nickel(II) complexes of generalformulae [Ni(L)

2] (1) [Ni(L)(B)] (2 3) where L is ligand

4m3BTSC (4-methoxy-3-benzyloxybenzaldehyde thiosemi-carbazide) and B is NN-donor heterocyclic base namely110-phenanthroline (phen) 221015840-bipyridine (bpy)

2 Experimental

21 Materials and Instrumentation All reagents and chemi-cals were of AR grade and used as purchasedThe 4-methoxy-3-benzyloxy benzaldehyde thiosemicarbazide and variousmetal salts were Merck products and used as supplied Theagarose (molecular biology grade) and ethidium bromide(EB) were obtained from Sigma Calf thymus (CT) DNAand Supercoiled (SC) pUC19 DNA (cesium chloride puri-fied) were purchased from Bangalore Genie (India) Thetris(hydroxymethyl)-aminomethane-HCl (tris-HCl) bufferwas prepared by doubly distilled H

2O

Bacterial Strains The four Salmonella typhimurium mutantstrains (histidine-dependant) TA98 TA100 TA1535 andTA1538 were procured from IMTECH Chandigarh IndiaFor mutagenicity assay each bacterial culture was inoculatedin 10mL of fresh nutrient broth and incubated at 37∘C for14 h All chemicals and bacterial media were purchased fromHimedia

The elemental analyses were carried out by using vario-micro CHNS15106062 analyzer 1HNMR and ESI-MS data ofthe compounds were recorded at the Sophisticated Analyticaland Instrument Facility IISc Bangalore The IR spectra ofthe samples were recorded on a Shimadzu spectrophotometerfrom 4000 to 400 cmminus1 using KBr pellets The UV-Vis spec-tra were recorded on a Shimadzu UV-3101PC spectropho-tometer using DMF as solvent The molar conductances of

the complexes were measured using Equiptronics digitalconductivitymeter number EQ-660A and themelting pointswere checked bymelting point apparatus used in laboratoriesMagnetic susceptibility data at 300K for the polycrystallinesamples of the complexes were obtained byVSM IITMadras

22 Synthesis The Schiff base ligand 4m3BTSC was synthe-sized by reported procedure [21]

221 Preparation of [Ni(L)2](X) (1) (L = 4m3BTSC X =DMF)

and [Ni(L)(B)] (2 3) (B = phen bpy) Complexes The com-plexes were prepared by following a modified reported [22]synthetic procedure in which a 10mL methanolic solutionof Ni(CH

3COO)

2sdot4H2O (024 g 1mmol) was reacted with

4m3BTSC (0315 g 1mmol) in acetonitrile (10mL) undermagnetic stirring at room temperatureThe resulting precipi-tate was filtered off washedwithmethanol and air-driedTheobtained precipitate was dissolved in dimethyl formamideand leaving it to stand for a week on slow evaporationof the solution at room temperature yielded a crystallinematerial (1) Crystals of complexes were suitable for X-raydiffraction studies After 30min a 20mLmethanolic solutionof the heterocyclic base [phen (0198 g 1mmol) bpy (0156 g1mmol)] was added to the solution and the resultingmixturewas stirred for 2 h at room temperature On cooling thesolution to an ambient temperature it was filtered and thefiltrate on slow concentration yielded a crystalline solid ofthe product (2 3) The solid was isolated and washed withcold methanol and finally dried over P

4O10 The FT-IR 1H

NMR and ESI-MS spectras for all the complexes are given insupplementary material (Figures S1ndashS9)

Analyses Calculaed for Complex 1 (C32H34N6NiO4S2) CHNS

Analyses Calc C 5574 H 499 N 1219 S 960 found C5313 H 505 N 1303 S 943 FT-IR cmminus1 (KBr disc)3420m (NH

2) 3180m (NndashH) 1596 s (C=N) 1522m (C=C)

1020m (ndashNndashN) 800m (C=S) 466m (NindashN) cmminus1 forcomplex 1 120582max nm (120576dm3Mminus1 cmminus1) in DMF 532(490)438(580) 370(1600) 340(2220) 285(1230) 268(23833) 120583eff(solid 300K) 041 120583B ΛM (Ωminus1 cm2Mminus1) in DMF at 25∘C117 ESI-MS 68801

Analyses Calculaed for Complex 2 (C28H25N5NiO2S) CHNS

Analyses Calc C 6067 H 455 N 1263 S 579 foundC 6132 H 488 N 1282 S 581 FT-IR cmminus1 (KBrdisc) 3420m (NH

2) 3180m (NndashH) 1600 s (C=N) 1522m

(C=C) 1020m (ndashNndashN) 817m (C=S) 455m (NindashN) cmminus1120582maxnm (120576dm3Mminus1 cmminus1) in DMF 530(470) 434(700)370(1550) 338(2520) 272(2530) (solid 300K) 030 120583B ΛM(Ωminus1 cm2Mminus1) in DMF at 25∘C 92 ESI-MS 5537



2) 3180m (NndashH) 1600 s (C=N) 1522m

(C=C) 1020m (ndashNndashN) 810m (C=S) 455m (NindashN) cmminus1120582maxnm (120576dm3Mminus1 cmminus1) in DMF 528(530) 435(860)

ISRN Inorganic Chemistry 3

375(2260) 338(2870) 268(2450) 120583eff (solid 300K) 040 120583BΛM (Ωminus1 cm2Mminus1) in DMF at 25∘C 84 ESI-MS 5296

23 X-Ray Crystallographic Procedures Single crystal of thecomplex 1 was grown by slow evaporation of DMF solutionDarkgrey single crystal was mounted on a glass fiber withepoxy cement The X-ray diffraction data were measured inframes with increasing 120596 (width of 03∘ per frame) and witha scan speed at 15 sframe on a Bruker SMART APEX CCDdiffractometer equippedwith a fine focus 175 kWsealed tubeX-ray source Empirical absorption corrections were carriedout using multiscan program [23] The structure was solvedby the heavy atom method and refined by full matrix least-squares using SHELX system of programs [24]

24 Antioxidant Activity

241 22-Diphenyl-1-picryl-hydrazyl (119863119875119875119867∙) Radical Scav-enging Method The reduction capability of DPPH radicalswas determined by the decrease in its absorbance at 517 nmwhich can be induced by antioxidants [25] Antioxidantactivity of ligand andmetal complexes was determined by themethod described by Brand-Williams et al [26]The complexsolutions (100120583L 1mmol) were mixed with 08mL of tris-HCl buffer (pH 74) to which 1mL of DPPH∙ (500120583M inethanol) was added The mixture was shaken vigorously andleft to stand for 30min Absorbance of the resulting solutionwas measured at 517 nm in a Uv-Vis SpectrophotometerThe radical scavenging activity was measured by decrease inthe absorbance of DPPH Lower absorbance of the reactionmixture indicated higher free radical scavenging activity

25 Antimicrobial Activity The antibacterial activity wastested against clinical isolates like Bacillus subtilis Micro-coccus luteus Staphylococcus aureus Klebsiella pneumoniaeEscherichia coli andPseudomonas aeruginosaThe test organ-ismsweremaintained on nutrient agar slants In vitro antibac-terial activity was determined by the agar well-diffusionmethod as described by Mukherjee et al [27] The overnightbacterial culture was centrifuged at 8000 rpm for 10minThebacterial cells were suspended in saline to make a suspensionof 105 CFUmL and used for the assay Plating was carried outby transferring the bacterial suspension to a sterile Petri plateand mixed with molten nutrient agar medium allowing themixture to solidify About 75120583L of the sample (2mgmL) wasplaced in the wells Plates were incubated at 37∘C and activitywas determined by measuring the diameter of the inhibitionzones The assay was carried out in triplicate

26 Mutagenicity Assay Mutagenicity of compounds wasstudied by the preincubation assay as described by Maronand Ames [28] 100120583L of fresh culture of Salmonella strainswas treated with test compounds dissolved in DMSO withconcentrations of 2 5 and 10mgplate at 37∘C For themutagenicity assay the controls and compound-treated cellswere mixed with 2mL of sterile top agar and poured ontominimal glucose agar plateThe plates were then inverted andincubated at 37∘C for 48 hrs Revertant colonies of bacteria

were countedmanually after 48 h Each experiment consistedof three replicates for each concentration

27 DNA Binding Experiments The UV absorbance at 260and 280 nm of the CT-DNA solution in 5mM tris-HCl buffer(pH 72) gave a ratio of 19 indicating that the DNA was freeof protein [29] The concentration of CT DNA was measuredfrom the band intensity at 260 nm with a known 120576 value(6600Mminus1 cmminus1) [30] Absorption titration measurementswere done by varying the concentration of CT DNA keepingthe metal complex concentration constant in 5mM tris-HCl5mM NaCl buffer (pH 72) Samples were kept forequilibrium before recording each spectrum The intrinsicbinding constant (119870

119887) for the interaction of the complexes

with CT-DNAwas determined from a plot of [DNA](120576119886minus120576

119891)

versus [DNA] using absorption spectral titration data and thefollowing equation

[DNA](120576

119886minus 120576

119891)

=

[DNA](120576

119887minus 120576

119891)

+ [K119887(120576

119887minus 120576

119891)]

minus1

(1)

where [DNA] is the concentration of DNA in base pairs Theapparent absorption coefficients 120576

119886 120576119891 and 120576

119887correspond to

119860obsd[Ni] the extinction coefficient for the free nickel(II)complex and extinction coefficient for the nickel(II) complexin the fully bound form respectively [31] 119870

119887is given by the

ratio of the slope to the interceptThe apparent binding constant (119870app) of the complexes

was determined by fluorescence spectral technique usingethidium bromide (EB) bound CT DNA solution in tris-HClNaCl buffer (pH 72) The fluorescence intensities of EBat 600 nm (546 nm excitation) with an increasing amount ofthe ternary complex concentration were recorded Ethidiumbromide was nonemissive in tris-buffer medium due to fluo-rescence quenching of the free EB by the solvent molecules[32 33] In the presence of DNA EB showed enhancedemission intensity due to its intercalative binding to DNAA competitive binding of the copper complexes to CT DNAcould result in the displacement of EB or quenching of thebound EB by the paramagnetic nickel(II) species decreasingits emission intensity

DNA-melting experiments were carried out by moni-toring the absorbance of CT-DNA (200120583M NP) at 260 nmwith varying temperature in the absence and presence of thecomplexes in 2 1 ratio of DNA to complex with a ramp rateof 05∘Cminminus1 in phosphate buffer (pH 685) using a Peltiersystem attached to the UV-Vis spectrophotometer

28 DNA Cleavage The extent of cleavage of supercoiled(SC) DNA in the presence of the complex and oxidizingagent H

2O2was determined by agarose gel electrophoresis

In a typical reaction supercoiled pUC19 DNA (02120583g) takenin 50mM tris-HCl buffer (pH 72) containing 50mM NaClwas treated with the complex The extent of cleavage wasmeasured from the intensities of the bands using UVITECGel Documentation System For mechanistic investigationsinhibition reactions were done on adding the reagents priorto the addition of the complex The solutions were incubated


Table 1 Analytical and physical data of the ligand and its complexes

Complex 1 2 3Colour aMP∘C Dark brown 190 Cream 195 Cream 195bIR (cmminus1)](C=N) + ](C=S) 1596 800 1600 817 1600 810cd-d band (nm) 532 530 530dΛMΩ

minus1cm2molminus1 117 92 84e120583eff 03 04 04

aMelting point bKBr Phase cin DMF medium din DMF medium at 25∘C and e120583eff for solid at 300K

for 1 h in a dark chamber at 37∘C followed by addition tothe loading buffer containing 25 bromophenol blue 025xylene cyanol and 30 glycerol (2 120583L) and the solutionwas finally loaded on 08 agarose gel containing 10 120583gmLethidium bromide (EB) Electrophoresis was carried out for2 h at 60V in tris-acetate-EDTA (TAE) buffer Bands werevisualized by UV light and photographed for analysis Duecorrections were made to the observed intensities for the lowlevel of NC form present in the original sample of SC DNAand for the low affinity of EB binding to SC in comparison tonicked-circular (NC) and linear forms of DNA [34]

3 Results and Discussion

31 Synthesis and General Aspects The new nickel(II) com-plexes are prepared in high yield from the reaction ofNi(CH

3COO)

2sdot4H2O with ligand 4m3BTSC and hetero-

cyclic bases The complexes were soluble in DMF DMSOand sparingly soluble in acetonitrile chloroform anddichloromethane The analytical data correspond well withthe general formula [ML

2] (DMF) (1) or [MLB] where M

is nickel L = 4m3BTSC and B is 110-phen 22-bpy (2 3)The proposed structures of complexes 2 and 3 are given inFigure 1 The molar conductance values of 1mM solutions ofthe metal complexes in DMF at room temperature are in therange of 8ndash12Ωminus1cm2molminus1 indicating their nonelectrolyticnature [35] They are characterized from the analytical andphysicochemical data (Table 1) The complexes 2 and 3display an intense charge transfer (CT) band in the range200ndash310 nmwhich can be attributed to the120587 rarr 120587

lowast transitionof the coordinated NN-donor heterocyclic base The bandaround 330 and 340 nm in all the complexes is due tothe imine (ndashHC=Nndash) and C=S groups and the occurrenceof two d-d bands in the regions 527ndash538 and 430ndash438 nm(1A1grarr1B

2g and 1A

1grarr1B

1g transitions) is consistent with

a square planar structure for all the all complexes

32 Crystal Structure of 1 The Ni(L)2(DMF) complex is

structurally characterized using single-crystal X-ray diffrac-tion techniqueThe ORTEP diagram and numbering schemefor Ni(L)

2are shown in Figure 2 Crystallographic data

for complex 1 is presented in Table 2 X-ray studies revealthat the complex has square planar geometry as is evidentfrom their bond parameters and bond angles (Table 3)Complex 1 crystallizes in the centrosymmetric P2

1n space

group of the monoclinic crystal system with cell parameters119886 = 15669(5) A 119887 = 9127(5) A 119888 = 16227(5) A and

Table 2 Crystallographic data for NiL2(1) complex

Formula C32H24N6NiO4S2Formula weight 68840Crystal system space group Monoclinic P2

1119899

119886 119887 119888(A) 15669(5) 9127(5) 16227(5)120572

∘ 120573∘ 120574∘ 90 1162(5) 90119881 (A3) 119885 20814(15) 4119879 (K) 293120579 238∘

120583 120mmminus1

120582 (A) (MoK120572) 071073119863

1199092168Mgmminus3

Refinement method Full-matrix least-squares on 1198652

Goodness of fit on 1198652 107119877int 0027Final 119877 indices 119908119877(119865

2) = 0134

Table 3 Selected bond lengths (A) and bond angles (∘) for NiL2(1)complex

Ni1ndashN1i 1937(2) N1ndashN2 1386(3)Ni1ndashN1 1937(2) N2ndashC16 1295(4)Ni1ndashS1 21614(11) N3ndashC16 1335(4)Ni1ndashS1i 21614(11) N4ndashC19 1323(7)S1ndashC16 1716(3) N4ndashC18 1384(7)N1indashNi1ndashN1 18000(9) C18ndashN4ndashC17 1220(7)N1indashNi1ndashS1 9426(8) O1ndashC6ndashC7 1083(3)N1ndashNi1ndashS1 8574(8) O1ndashC8ndashC9 1247(3)N1indashNi1ndashS1i 8574(8) O1ndashC8ndashC13 1146(3)N1ndashNi1ndashS1i 9426(8) C9ndashC8ndashC13 1207(3)S1ndashNi1ndashS1i 18000(4) C8ndashC9ndashC10 1203(3)C16ndashS1ndashNi1 9715(11) O2ndashC13ndashC12 1254(3)C8ndashO1ndashC6 1172(2) O2ndashC13ndashC8 1150(3)C13ndashO2ndashC14 1180(2) C12ndashC13ndashC8 1195(3)C15ndashN1ndashN2 1142(2) N1ndashC15ndashC10 1334(3)C15ndashN1ndashNi1 1258(2) N2ndashC16ndashN3 1196(3)N2ndashN1ndashNi1 11996(19) N2ndashC16ndashS1 1228(2)C16ndashN2ndashN1 1139(2) N3ndashC16ndashS1 1176(2)C19ndashN4ndashC18 1193(7) O2ndashC13ndashC12 1254(3)C19ndashN4ndashC17 1187(5) O2ndashC13ndashC8 1150(3)

119881 = 20814(15) A3 All nonhydrogen atoms were refinedanisotropically and the hydrogen was refined isotropically


N

O

ONi

HN

S

NN

N

OO

N N

HN

S

Ni

(2) (3)

NH2 NH2

Figure 1 Proposed structures of Ni(II) complexes 2 and 3

Figure 2 The ORTEP and ball and stick view of the complex Ni(L)2(DMF) (1)

The hydrogen atoms attached to the heteroatoms were intheir calculated positions and refined according to the ridingmodel The coordination polyhedron around the nickel atomis best described as square planar with an N

2S2donor set

The nickel atom is located at the centre of symmetry andis coordinated by two sulphur atoms and two azomethinenitrogen atoms from two Schiff base ligands in usual transarrangementThe trans bond angle ofN(1)ndashNindashN(1) is exactly180∘ and the bond angle between S(1)ndashNindashS(1) is also 180∘this confirms that structure is square planar The crystalpacking viewed along the cell axis displays nine independentmolecules accommodated in the crystallographic asymmet-ric unit in which eight molecules occupy eight corners of theunit cell and one more molecule is occupied in the centre ofthe unit cell The structure shows extensive intermolecularnoncovalent interactions and it is also stabilized byNndashHsdot sdot sdotOintermolecular hydrogen bonding Here the atom N3 acts asa donor to O atom of the lattice DMF molecule (distance2678 A) The unit cell packing diagram present in complex1 is shown in Figure 3(a) and the intermolecular hydrogenbonding is shown in Figure 3(b)

33 IR Spectra In principle the ligand can exhibit thione-thiol tautomerism since it contains a thioamide ndashNHndashC=Sfunctional group The ](SndashH) band at 2565 cmminus1 is absent inthe IR spectrum of ligand but ](NndashH) band at sim3265 cmminus1 ispresent indicating that in the solid state the ligand remainsas the thione tautomer The position of ](C=N) band of thethiosemicarbazone appeared at 1616 cmminus1 is shifted towardslower wave number in the complexes indicating coordinationvia the azomethine nitrogen This is also confirmed by

the appearance of band in complexes in the range of 450ndash455 cmminus1 which has been assigned to the ](MndashN) [36] Amedium band found at 1014 cmminus1 is due to the ](NndashN) groupof the thiosemicarbazoneThe position of this band is shiftedtowards higher wave number in the spectra of complexesIt is due to the increase in the bond strength which againconfirms the coordination via the azomethine nitrogen Theband appearing at sim830 cmminus1 corresponding to ](C=S) in theIR spectrum of ligand is shifted towards lower wave numberIt indicates that thione sulfur coordinates to the metal ion[37] Thus it may be concluded that the ligand behaves asbidentate chelating agent coordinating through azomethinenitrogen and thiolate sulfurTheCndashH stretching and bendingvibrations appear at 2780ndash2890 cmminus1 and 1460ndash1420 cmminus1respectively

34 1H NMR Spectra The 1H NMR spectra of ligand andits nickel complexes were recorded in CDCl

3 The 1H NMR

spectrum of 4m3BTSC shows signal at 830 120575 indicating thepresence of azomethine (HC=N) proton The singlet peaksbetween 93ndash101 120575 and sim68 120575 are assigned for the presenceof NH and NH

2protons however the multiplet in the region

710ndash76 120575 indicates the presence of the phenolic protons Nosignals are observed in the range 113-114 120575 which indicatesthe absence of thiol (ndashSH) group The other importantbands were observed at sim39 120575 assigned for methyl (OndashCH

3)

protons and 516 120575 assigned for methylene (OndashCH2) protons

The azomethine proton in the Ni(II) complexes is shifteddownfield compared to free ligand suggesting deshieldingof the azomethine group due to coordination with nickel


(a) (b)

Figure 3 (a) Molecular packing for the complex NiL2 viewed down the c-axis (b) dashed lines indicate hydrogen bonds

Table 4 1H NMR spectra of ligand and its complexes

Compounds ndashNH (120575) ndashCH=Nndash (120575) ndashPhndash (120575) ndashNH2 (120575) ndashCH2 ndashOCH3 (120575)

4m3BTSC 1007 (s 1H NH) 830 (s 1H) 71ndash74 (m) 802 (s 2H NH2)516 (s 2H CH2)390 (s 3H CH3)

1 101 (s 1H NH) 781 (s 1H) 72ndash74 (m) 688 (s 2H NH2)516 (s 2H CH2)390 (s 3H CH3)



s singlet and m multiplet

Table 5 DPPH radical scavenging activity of complexes 1 2 and 3and standards

Compounds Conc (120583M) aEC50bARP cAE

4m3BTSC mdash NI NI NI1 40 0110 62 347 times 10minus3

2 50 0110 50 27 times 10minus3

3 50 0105 52 29 times 10minus3

Vit-C 42 0168 83 46 times 10minus3

BHT 20 008 125 69 times 10minus3

NI no inhibition aEC50 effective concentration = 120583M conc of antioxi-dant120583M conc of DPPHbARP antioxidant radical power = 1EC50cAE antioxidant efficiency = 1EC50 119879EC50 Vit-C vitamin CBHT butylated hydroxy anisole

There is no appreciable change in the other signals of thecomplexes (Table 4)


351 DPPH Radical Scavenging Activity Various researchershave used scavenging effect of a chemical on DPPH radical asa quick and reliable parameter to assess the in vitro antioxi-dant activityThe efficient concentration of complex required

to scavenge 50 of DPPH EC50 is shown in Table 5 The

lower the EC50is the better its radical scavenging activity is

It is evident from results that free radical scavenging activitiesof these compounds were concentration dependent Amongthe examined compounds the complex 1 showed a stronginteractive ability with DPPH expressed as EC

50value of

40 120583Mconcentration which was close to that of ascorbic acid(30 120583M) Maximum free radical scavenging activity (569)was found in complex 1 followed by complex 3 (5241) andcomplex 2 (518) while least activity (3288) was observedfor free ligand 4m3BTSC The comparative time-dependentantioxidant activity of compounds and standards were shownby graph in Figure 4 Antiradical efficiency (AE) involves thepotency (1EC

50) and the reaction time (119879EC

50

) The AE of allthe complexes are in the range of 25ndash35times 10minus3 the value nearto standards and the complexes are good antioxidants [38]

36 Antibacterial Activity The in vitro antibacterial activityof the Schiff bases and their Ni(II) complexes were evaluatedagainst three Gram positive S aureus B subtilis and Mluteus and three Gram negative bacteria P aeruginosaK pneumoniae and E coli Table 6 illustrates the antimi-crobial activity of the synthesized compounds In generalthe synthesized Schiff bases have more antibacterial activitycompared to their metal complexes this may be due to


Table 6The values of zone inhibition (mm) of microorganisms forthe 4m3BTSC and its complexes 1 2 and 3

Bacteria 4m3BTSC 1 2 3E coli(MTCC 443) 10 10 12 mdash

P aeruginosa(MTCC 741) mdash mdash 11 mdash

K pneumonia(MTCC 109) 12 mdash 14 10

B subtilis(MTCC 441) 12 10 15 9

M luteus(MTCC 106) 11 mdash 12 mdash

S aureus(MTCC 3160) mdash mdash 12 mdash

the greater liphophilic nature of the Schiff bases than theirmetal complexes The results showed that the complex 2exhibited antibacterial activity against all the tested organ-isms namely E coli P aeruginosa K pneumoniae B subtilisM luteus and S aureus The result also indicated that thecomplex 1 exhibited antibacterial activity against E coli andB subtilis and that complex 3 exhibited antibacterial activitiesagainst K pneumonia and B subtilis respectivelyThe factorsthat govern antimicrobial activities are strongly dependenton the central metal ion and coordination numbers and arealso due to the presence of nitrogen and sulfur donor groups[39 40]

37 Mutagenic Assay Themutagenic activity was carried outby Ames test The results indicate that all the complexesare nonmutagenic in nature under assay condition Accord-ing to the Ames test the tested ligand and its complexesdid not show mutagenicity with bacterial strain Salmonellatyphimurium types TA 98 TA 100 TA 1535 and TA 1538 inthe assayed range

38 DNA Binding Properties It was reported that DNAbindingmode and affinity are affected by a number of factorssuch as planarity of ligands [41] the coordination geometrythe ligand donor atom type [42] the metal ion type and itsflexible valence [43] The UV binding of the complexes 1ndash3to the calf thymus (CT) DNA has been studied by electronicabsorption spectral technique

The intrinsic binding constant (119870119887) value was 399(plusmn004)

times 106Mminus1 for complex 2 (Figure 5)The complexes 1 and 3 didnot show binding affinity to CT-DNA due to lack of planarityThe experimental values of119870

119887revealed that complex 2 binds

to DNA via intercalative mode and this may be due to thepresence of the phenanthroline ring in the structure

The emission spectral method is used to study the relativebinding of the complexes to CT-DNAThe emission intensityof ethidium bromide (EB) is used as a spectral probe EBshows reduced emission intensity in buffer solution becauseof solvent quenching and an enhancement of the emissionintensity when intercalatively bound to DNAThe binding of

0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time

LigandNiLXNiLPhenNiLBPY

DPPHBHTVitC

Figure 4 The comparative time-dependent antioxidant activity ofcomplexes 1 2 and 3 and standards

250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)

Figure 5 (a) Absorption spectral traces showing the decrease inabsorption intensity on gradual addition of CT-DNA (270120583M) inaliquots to the solution of complex 2 (60120583M) in 5mM tris-HClbuffer (pH 72) at 25∘C (shown by arrow) (b) Inset shows plot of[DNA](120576

119886minus 120576

119891) versus [DNA] for absorption titration of CT-DNA

with complex 2

the complexes to DNA decreases the emission intensity of EB(Figures 6(a) 6(b) and 6(c))The relative binding propensityof the complexes to DNA is measured from the extent ofreduction in the emission intensity (Figure 7) The apparentbinding constant (119870app) values for 1 2 and 3 are given inTable 7 The phen complex showed significantly higher 119870appvalues than those of the bpy and bis ligand complexesThe bisligand complex is a poor binder to DNA

The nature of the binding of the complexes to CT-DNA was further investigated by DNA melting experimentThermal behavior of the DNA in the presence of metal


580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)

Figure 6The emission spectral changes on addition of complexes 1 (a) 2 (b) and 3 (c) to the CT-DNA bound to ethidium bromide (shownby arrow)

0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0

Complex 1 Kapp = 17 times 105 mminus1



Figure 7 The effect of addition of metal complexes 1 2 and 3 tothe emission intensity CT-DNAbound ethidium bromide in a 5mMtris-HCl (pH = 72) at 25∘C

complexes can give insight into their conformational changesand information about the interaction strength with DNADNAmelting is observed when dsDNAmolecules are heated

Table 7 Intrinsic binding apparent DNA binding and meltingtemperature studies

Complexes a119870

119887Mminus1 b

119870appMminus1 cΔ119879

119898

∘C1 17 times 105 sdot sdot sdot

2 399 times 106 1098 times 105 +083 319 times 105 minus04aIntrinsic binding constant from absorption spectral methodbApparent binding constant from competitive binding assay by emissionmethodcChanges in melting temperature of CT-DNA

and separated into two single strands it occurs due toa disruption of the intermolecular forces such as 120587 stackingandhydrogen bonding interactions betweenDNAbase pairs

A classical intercalator such as EB stabilizes the duplexDNA causing the DNA to melt at a higher temperature [44]The melting temperature of CT-DNA in the absence of anyadded complex was found to be 655∘C in our experimentalconditions Under the same set of conditions in the presenceof all the complexes they gaveΔ119879

119898values ranging from04 to

+08∘C indicating primarily DNA groove binding propensityof the complexes The phen complex 2 with a relativelyhigh Δ119879

119898value of +08∘C could have a partial intercalative


Table 8 Selected cleavage data of SC pUC19 by ligand and itscomplexes 1 2 and 3

Lane no Conditions [Complex] 120583M aSC bNC1 DNA control mdash 97 32 DNA + Ligand + H2O2 60 92 83 DNA + Ligand + MPA 60 93 74 DNA + 1 + MPA 60 90 105 DNA + 2 + MPA 60 87 136 DNA + 3 + MPA 60 89 117 DNA + 1 + H2O2 60 78 228 DNA + 2 + H2O2 60 65 359 DNA + 3 + H2O2 60 75 25aSC supercoiled DNAbNC nicked circular DNA

mode of DNA binding with the planar ring of 110-phenintercalating to the DNA base pairs

39 DNA Cleavage Studies The oxidative DNA cleavageactivity of the complexes in the presence of oxidizing agenthydrogen peroxide (H

2O2 50mM) is investigated by agarose

gel electrophoresis using supercoiled (SC) plasmid pUC19DNA (02120583g 333 120583M NP) in 50mM tris-HCl50mM NaClbuffer (pH 72) and the selected DNA cleavage data are givenin Table 8 The phen complex 2 (60 120583M) shows moderateldquochemical nucleaserdquo activity and 35 conversion of the SC(form I) to its nicked-circular form (NC form II) of DNA isobserved Control experiments with H

2O2or the complexes

alone did not show any apparent conversion of SC to itsnicked-circular (NC) form (Figure 8)The bpy complex 3 andbis ligand complex 1 are inactive due to their poor bindingability to DNA The pathways involved in the DNA cleavagereactions are believed to be analogous to those proposed bySigman and coworkers for the chemical nuclease activity ofbis(phen)copper species (Scheme 1) [45 46] Based on themechanistic studies it is observed that chemical nucleaseactivity is high in oxidizing condition compared to reducingcondition

4 Conclusions

Three new Ni(II) complexes having NS-donor ligand andNN-donor heterocyclic bases are prepared and character-ized The complex 1 was structurally characterized by X-raycrystallography and the structure shows the square planargeometry The free radical scavenging assay results showedthat complex 1 posseses significant activity when comparedto complexes 2 and 3 The biological studies showed that theligand and its complexes exhibited significant and differentbiological activities and also the prepared compounds arenonmutagenic They may be potential commercial antioxi-dants because of their nonmutagenic and nontoxic natureThe DNA interaction of the new complexes was evaluated byabsorption emission andmelting temperaturemethods andthe results suggested that the binding affinity of the complexesincreases with the presence of planar ligand in the molecule

NiII(L)2(LB) + DNA NiII(L)2

Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙

Scheme 1 Proposed mechanistic pathway for the chemical nucleaseactivity of complexes in tris-buffer medium

Lane 3 6 2 1 4 5 7 8 9

NC

SC

Figure 8 Gel electrophoresis diagram showing the cleavage of SCpUC19 DNA by complexes 1 2 and 3 and ligand in 50mm tris-HCL50mm NaCl buffer (pH 72) in the presence oxidizing andreducing agents

The nickel(II) complexes with planar phenanthroline basesshow moderate DNA binding and cleavage ability

Acknowledgments

The authors gratefully acknowledge the support of ProfessorAkhil R Chakravarty Department of Inorganic and Physicalchemistry Indian Institute of Science Bangalore India inproviding facilities and useful discussions Financial supportreceived from University Grants Commission (UGC Ref noF No 39-7542010(SR) dated 13-01-2011) India is gratefullyacknowledged M N Somashekar is thankful to SCSTcell and DST-PURSE Programme Bangalore University forfellowship The authors are also thankful to the UniversitiesSophisticated Instrumental Centre Karnataka UniversityDharwad for the X-ray data collection

References

[1] N Raman S Johnson Raja and A Sakthivel ldquoTransition metalcomplexes with Schiff-base ligands 4-Aminoantipyrine basedderivativesmdasha reviewrdquo Journal of Coordination Chemistry vol62 no 5 pp 691ndash709 2009

[2] M A Ali N E H Ibrahim R J Butcher J P Jasinski JM Jasinski and J C Bryan ldquoSynthesis and characterizationof some four-and five-coordinate copper(II) complexes of 6-methyl-2-formylpyridinethiosemicarbazone (HNNS) and theX-ray crystal structures of the [Cu(NNS)(CH

3COO)(H

2O)]

and [Cu(HNNS)(H2O)(SO

4)]sdotH2O complexesrdquo Polyhedron

vol 17 no 11-12 pp 1803ndash1809 1998[3] Y-J Kim S-O Kim Y-I Kim and S-N Choi ldquoA mimic

molecule of blue copper protein active site [(ndash)-sparteine-NN1015840](maleonitriledithiolato-SS1015840)copper(II)rdquo Inorganic Chem-istry vol 40 no 17 pp 4481ndash4484 2001

[4] M M van Dyke and P B Dervan ldquoEchinomycin binding siteson DNArdquo Science vol 225 no 4667 pp 1122ndash1127 1984


[5] B G Patil B R Havinale J M Shallom and M P ChitnisldquoSyntheses and spectroscopic studies of potential antitumorcopper(II) complexeswith 5-phenylazo-3-methoxy salicylidenethiosemicarbazone and N4 substituted thiosemicarbazonesrdquoJournal of Inorganic Biochemistry vol 36 no 2 pp 107ndash113 1989

[6] P V Bernhardt J Mattsson and D R Richardson ldquoComplexesof cytotoxic chelators from the dipyridyl ketone isonicotinoylhydrazone (HPKIH) analoguesrdquo Inorganic Chemistry vol 45no 2 pp 752ndash760 2006

[7] M Akbar Ali and S E Livingstone ldquoMetal complexes ofsulphur-nitrogen chelating agentsrdquo Coordination ChemistryReviews vol 13 no 2-3 pp 101ndash132 1974

[8] C Costello T Karpanen P A Lambert et al ldquoThiosemicar-bazones active against Clostridium difficilerdquo Bioorganic andMedicinal Chemistry Letters vol 18 no 5 pp 1708ndash1711 2008

[9] J S Sebolt S V Scavone and C D Pinter ldquoPyrazoloacridinesa new class of anticancer agents with selectivity against solidtumors in vitrordquoCancer Research vol 47 no 16 pp 4299ndash43041987

[10] S Rekha and K R Nagasundara ldquoComplexes of the Schiffbase derived from 4-aminophenyl benzimidazole and 221015840-dehydropyrollidene-N-aldehyde with Zn (II) Cd (II) and Hg(II) halidesrdquo Indian Journal of Chemistry A vol 45 no 11 pp2421ndash2425 2006

[11] D Kovala-Demertzi A Papageorgiou L Papathanasis et alldquoIn vitro and in vivo antitumor activity of platinum(II) com-plexes with thiosemicarbazones derived from 2-formyl and2-acetyl pyridine and containing ring incorporated at N(4)-position synthesis spectroscopic study and crystal structureof platinum(II) complexes with thiosemicarbazones potentialanticancer agentsrdquo European Journal of Medicinal Chemistryvol 44 no 3 pp 1296ndash1302 2009

[12] M C Rodrıguez-Arguelles M B Ferrari F Bisceglie et alldquoSynthesis characterization and biological activity of Ni Cuand Zn complexes of isatin hydrazonesrdquo Journal of InorganicBiochemistry vol 98 no 2 pp 313ndash321 2004

[13] F Liang P Wang X Zhou et al ldquoNickle(II) and cobalt(II)complexes of hydroxyl-substituted triazamacrocyclic ligand aspotential antitumor agentsrdquo Bioorganic and Medicinal Chem-istry Letters vol 14 no 8 pp 1901ndash1904 2004

[14] J Suh ldquoSynthetic artificial peptidases and nucleases usingmacromolecular catalytic systemsrdquo Accounts of ChemicalResearch vol 36 no 7 pp 562ndash570 2003

[15] R T Kovacic J TWelch and S J Franklin ldquoSequence-selectiveDNA cleavage by a chimeric metallopeptiderdquo Journal of theAmerican Chemical Society vol 125 no 22 pp 6656ndash66622003

[16] D S Sigman ldquoNuclease activity of 110-phenanthroline-copperionrdquo Accounts of Chemical Research vol 19 no 6 pp 180ndash1861986

[17] D S Sigman T W Bruice A Mazumder and C L SuttonldquoTargeted chemical nucleasesrdquo Accounts of Chemical Researchvol 26 no 3 pp 98ndash104 1993

[18] K E Erkkila D T Odom and J K Barton ldquoRecognition andreaction of metallointercalators with DNArdquo Chemical Reviewsvol 99 no 9 pp 2777ndash2795 1999

[19] R M Burger ldquoCleavage of nucleic acids by bleomycinrdquo Chemi-cal Reviews vol 98 no 3 pp 1153ndash1169 1998

[20] B M Zeglis V C Pierre and J K Barton ldquoMetallo-intercalators and metallo-insertorsrdquo Chemical Communica-tions no 44 pp 4565ndash4579 2007

[21] M A S Aslam S-U Mahmood M Shahid A Saeed andJ Iqbal ldquoSynthesis biological assay in vitro and moleculardocking studies of new Schiff base derivatives as potentialurease inhibitorsrdquoEuropean Journal ofMedicinal Chemistry vol46 no 11 pp 5473ndash5479 2011

[22] R Rao A K Patra and P R Chetana ldquoSynthesis structureDNA binding and oxidative cleavage activity of ternary (l-leucineisoleucine) copper(II) complexes of heterocyclic basesrdquoPolyhedron vol 27 no 5 pp 1343ndash1352 2008

[23] N Walker and D Stuart ldquoAn empirical method for correctingdiffractometer data for absorption effectsrdquoActa Crystallograph-ica A vol 39 no 1 pp 158ndash166 1983

[24] G M Sheldrick SHELX-97 ldquoProgram For Crystal StructureSolution and Refinementrdquo University of Gottingen GottingenGermany 1997

[25] B Matthaus ldquoAntioxidant activity of extracts obtained fromresidues of different oilseedsrdquo Journal of Agricultural and FoodChemistry vol 50 no 12 pp 3444ndash3452 2002

[26] W Brand-Williams M E Cuvelier and C Berset ldquoUse of a freeradical method to evaluate antioxidant activityrdquo Lebensmittel-Wissenschaft amp Technologie vol 28 no 1 pp 25ndash30 1995

[27] P K Mukherjee R Balasubramanian K Saha B P Sahaand M Pal ldquoAntibacterial efficiency of Nelumbo nucifera(nymphaeaceae) rhizomes extractrdquo Indian Drugs vol 32 no 6pp 274ndash276 1995

[28] D M Maron and B N Ames ldquoRevised methods for theSalmonella mutagenicity testrdquo Mutation Research vol 113 no3-4 pp 173ndash215 1983

[29] J Marmur ldquoA procedure for the isolation of deoxyribonucleicacid from micro-organismsrdquo Journal of Molecular Biology vol3 pp 208ndash218 1961

[30] M E Reichmann S A Rice C A Thomas and P DotyldquoA further examination of the molecular weight and size ofdesoxypentose nucleic acidrdquo Journal of the American ChemicalSociety vol 76 no 11 pp 3047ndash3053 1954

[31] A Wolfe G H Shimer Jr and T Meehan ldquoPolycyclic aromatichydrocarbons physically intercalate into duplex regions ofdenatured DNArdquo Biochemistry vol 26 no 20 pp 6392ndash63961987

[32] J B LePecq and C Paoletti ldquoA fluorescent complex betweenethidium bromide And nucleic acids Physical-chemical char-acterizationrdquo Journal of Molecular Biolology vol 27 pp 87ndash1061967

[33] S Neidle ldquoDNAminor-grooverecognition by small moleculesrdquoNatural Product Reports vol 18 pp 291ndash309 2001

[34] J Bernadou G Pratviel F Bennis M Girardet and BMeunierldquoPotassium monopersulfate and a water-soluble manganeseporphyrin complex [Mn(TMPyP)](OAc)5 as an efficientreagent for the oxidative cleavage of DNArdquo Biochemistry vol28 no 18 pp 7268ndash7275 1989

[35] W J Geary ldquoThe use of conductivity measurements in organicsolvents for the characterisation of coordination compoundsrdquoCoordination Chemistry Reviews vol 7 no 1 pp 81ndash122 1971

[36] S Chandra and U Kumar ldquoSpectral studies of coordinationcompounds of cobalt(II) with thiosemicarbazone of hetero-cyclic ketonerdquo Spectrochimica Acta A vol 62 no 4-5 pp 940ndash944 2005

[37] N S Youssef and K H Hegab ldquoSynthesis and characterizationof some transition metal complexes of thiosemicarbazonesderived from 2-acetylpyrrole and 2-acetylfuranrdquo Synthesis andReactivity in Inorganic Metal-Organic and Nano-Metal Chem-istry vol 35 no 5 pp 391ndash399 2005


[38] C Sanchez-Moreno J A Larrauri and F Saura-Calixto ldquoAprocedure tomeasure the antiradical efficiency of polyphenolsrdquoJournal of the Science of Food and Agriculture vol 76 no 2 pp270ndash276 1998

[39] A R Burkanudeen R S Azarudeen M A R Ahamedand W B Gurnule ldquoKinetics of thermal decomposition andantimicrobial screening of terpolymer resinsrdquo Polymer Bulletinvol 67 no 8 pp 1553ndash1568 2011

[40] M S Refat and N M El-Metwaly ldquoSpectral thermal andbiological studies of Mn(II) and Cu(II) complexes with twothiosemicarbazide derivativesrdquo Spectrochimica Acta A vol 92pp 336ndash346 2012

[41] C V Kumar J K Barton and N J Turro ldquoPhotophysics ofruthenium complexes bound to double helical DNArdquo Journalof the American Chemical Society vol 107 no 19 pp 5518ndash55231985

[42] S Mahadevan and M Palaniandavar ldquoSpectroscopic andvoltammetric studies of copper (II) complexes of bis(pyrid-2-yl)-ditrithia ligands bound to calf thymus DNArdquo InorganicaChimica Acta vol 254 no 2 pp 291ndash302 1997

[43] MAsadi E Safaei B Ranjbar and LHasani ldquoThermodynamicand spectroscopic study on the binding of cationic ZD(II) andCo(II) tetrapyridinoporphyrazines to calf thymusDNA the roleof the central metal in binding parametersrdquo New Journal ofChemistry vol 28 no 10 pp 1227ndash1234 2004

[44] J M Kelly A B Tossi D J Mcconnell and C Ohuigin ldquoAstudy of the interactions of some polypyridylruthenium(II)complexes with DNA using fluorescence spectroscopy topoiso-merisation and thermal denaturationrdquo Nucleic Acids Researchvol 13 no 17 pp 6017ndash6034 1985

[45] O Zelenko J Gallagher and D S Sigman ldquoScission of DNAwith Bis(110-phenanthroline)copper without IntramolecularHydrogen Migrationrdquo Angewandte Chemie (International Edi-tion) vol 36 no 24 pp 2776ndash2778 1997

[46] D S Sigman ldquoChemical nucleasesrdquo Biochemistry vol 29 no39 pp 9097ndash9105 1990

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


screened for inhibition of cancer cell proliferation [12 13]In the last 20 years transition metal complexes have becomeincreasingly important as drugs and artificial nucleasesOne goal of designing Ni(II) complexes is developing morereactive chemical systems that can efficiently cleave DNAunder physiological conditions [14ndash16]

Substantial progress has been made during the past fewdecades to develop metal-based small molecules as DNAfoot-printing as well as the therapeutic agents that are capableof binding and cleaving DNA under physiological condition[17 18] Among the metal complexes so far investigatedthose of polypyridyl phenanthroline bases have attractedgreat attention by virtue of its binding propensity to nucleicacid under the physiological condition [19 20]

In view of the varying ligation behavior and interestingbiological activity shown by these nitrogen-sulfur ligandsand their complexes we are also interested to preparethe thiosemicarbazide Schiff base and its metal complexesDesigning of these metal complexes has been undertakenand their biological activities have been studiedThe study ofbiological activities of benzyloxy benzaldehyde-Schiff basesis less explored Herein we present the syntheses struc-ture antioxidant antimicrobial and mutagenic activitiesand DNA interactions of nickel(II) complexes of generalformulae [Ni(L)

2] (1) [Ni(L)(B)] (2 3) where L is ligand

4m3BTSC (4-methoxy-3-benzyloxybenzaldehyde thiosemi-carbazide) and B is NN-donor heterocyclic base namely110-phenanthroline (phen) 221015840-bipyridine (bpy)

2 Experimental

21 Materials and Instrumentation All reagents and chemi-cals were of AR grade and used as purchasedThe 4-methoxy-3-benzyloxy benzaldehyde thiosemicarbazide and variousmetal salts were Merck products and used as supplied Theagarose (molecular biology grade) and ethidium bromide(EB) were obtained from Sigma Calf thymus (CT) DNAand Supercoiled (SC) pUC19 DNA (cesium chloride puri-fied) were purchased from Bangalore Genie (India) Thetris(hydroxymethyl)-aminomethane-HCl (tris-HCl) bufferwas prepared by doubly distilled H

2O

Bacterial Strains The four Salmonella typhimurium mutantstrains (histidine-dependant) TA98 TA100 TA1535 andTA1538 were procured from IMTECH Chandigarh IndiaFor mutagenicity assay each bacterial culture was inoculatedin 10mL of fresh nutrient broth and incubated at 37∘C for14 h All chemicals and bacterial media were purchased fromHimedia

The elemental analyses were carried out by using vario-micro CHNS15106062 analyzer 1HNMR and ESI-MS data ofthe compounds were recorded at the Sophisticated Analyticaland Instrument Facility IISc Bangalore The IR spectra ofthe samples were recorded on a Shimadzu spectrophotometerfrom 4000 to 400 cmminus1 using KBr pellets The UV-Vis spec-tra were recorded on a Shimadzu UV-3101PC spectropho-tometer using DMF as solvent The molar conductances of

the complexes were measured using Equiptronics digitalconductivitymeter number EQ-660A and themelting pointswere checked bymelting point apparatus used in laboratoriesMagnetic susceptibility data at 300K for the polycrystallinesamples of the complexes were obtained byVSM IITMadras

22 Synthesis The Schiff base ligand 4m3BTSC was synthe-sized by reported procedure [21]

221 Preparation of [Ni(L)2](X) (1) (L = 4m3BTSC X =DMF)

and [Ni(L)(B)] (2 3) (B = phen bpy) Complexes The com-plexes were prepared by following a modified reported [22]synthetic procedure in which a 10mL methanolic solutionof Ni(CH

3COO)

2sdot4H2O (024 g 1mmol) was reacted with

4m3BTSC (0315 g 1mmol) in acetonitrile (10mL) undermagnetic stirring at room temperatureThe resulting precipi-tate was filtered off washedwithmethanol and air-driedTheobtained precipitate was dissolved in dimethyl formamideand leaving it to stand for a week on slow evaporationof the solution at room temperature yielded a crystallinematerial (1) Crystals of complexes were suitable for X-raydiffraction studies After 30min a 20mLmethanolic solutionof the heterocyclic base [phen (0198 g 1mmol) bpy (0156 g1mmol)] was added to the solution and the resultingmixturewas stirred for 2 h at room temperature On cooling thesolution to an ambient temperature it was filtered and thefiltrate on slow concentration yielded a crystalline solid ofthe product (2 3) The solid was isolated and washed withcold methanol and finally dried over P

4O10 The FT-IR 1H

NMR and ESI-MS spectras for all the complexes are given insupplementary material (Figures S1ndashS9)

Analyses Calculaed for Complex 1 (C32H34N6NiO4S2) CHNS

Analyses Calc C 5574 H 499 N 1219 S 960 found C5313 H 505 N 1303 S 943 FT-IR cmminus1 (KBr disc)3420m (NH

2) 3180m (NndashH) 1596 s (C=N) 1522m (C=C)

1020m (ndashNndashN) 800m (C=S) 466m (NindashN) cmminus1 forcomplex 1 120582max nm (120576dm3Mminus1 cmminus1) in DMF 532(490)438(580) 370(1600) 340(2220) 285(1230) 268(23833) 120583eff(solid 300K) 041 120583B ΛM (Ωminus1 cm2Mminus1) in DMF at 25∘C117 ESI-MS 68801



2) 3180m (NndashH) 1600 s (C=N) 1522m

(C=C) 1020m (ndashNndashN) 817m (C=S) 455m (NindashN) cmminus1120582maxnm (120576dm3Mminus1 cmminus1) in DMF 530(470) 434(700)370(1550) 338(2520) 272(2530) (solid 300K) 030 120583B ΛM(Ωminus1 cm2Mminus1) in DMF at 25∘C 92 ESI-MS 5537



2) 3180m (NndashH) 1600 s (C=N) 1522m

(C=C) 1020m (ndashNndashN) 810m (C=S) 455m (NindashN) cmminus1120582maxnm (120576dm3Mminus1 cmminus1) in DMF 528(530) 435(860)












119891)


[DNA](120576

119886minus 120576

119891)

=

[DNA](120576

119887minus 120576

119891)

+ [K119887(120576

119887minus 120576

119891)]

minus1

(1)


119886 120576119891 and 120576

119887correspond to

















3COO)






2g and 1A

1grarr1B







1n space




1119899

119886 119887 119888(A) 15669(5) 9127(5) 16227(5)120572

∘ 120573∘ 120574∘ 90 1162(5) 90119881 (A3) 119885 20814(15) 4119879 (K) 293120579 238∘

120583 120mmminus1

120582 (A) (MoK120572) 071073119863

1199092168Mgmminus3



2) = 0134





N

O

ONi

HN

S

NN

N

OO

N N

HN

S

Ni

(2) (3)

NH2 NH2




2S2donor set





3 The 1H NMR




3)




(a) (b)












2 50 0110 50 27 times 10minus3

3 50 0105 52 29 times 10minus3










50value of



50



















0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time


DPPHBHTVitC


250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)


119886minus 120576


with complex 2




580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of












119891)


[DNA](120576

119886minus 120576

119891)

=

[DNA](120576

119887minus 120576

119891)

+ [K119887(120576

119887minus 120576

119891)]

minus1

(1)


119886 120576119891 and 120576

119887correspond to

















3COO)






2g and 1A

1grarr1B







1n space




1119899

119886 119887 119888(A) 15669(5) 9127(5) 16227(5)120572

∘ 120573∘ 120574∘ 90 1162(5) 90119881 (A3) 119885 20814(15) 4119879 (K) 293120579 238∘

120583 120mmminus1

120582 (A) (MoK120572) 071073119863

1199092168Mgmminus3



2) = 0134





N

O

ONi

HN

S

NN

N

OO

N N

HN

S

Ni

(2) (3)

NH2 NH2




2S2donor set





3 The 1H NMR




3)




(a) (b)












2 50 0110 50 27 times 10minus3

3 50 0105 52 29 times 10minus3










50value of



50



















0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time


DPPHBHTVitC


250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)


119886minus 120576


with complex 2




580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of









3COO)






2g and 1A

1grarr1B







1n space




1119899

119886 119887 119888(A) 15669(5) 9127(5) 16227(5)120572

∘ 120573∘ 120574∘ 90 1162(5) 90119881 (A3) 119885 20814(15) 4119879 (K) 293120579 238∘

120583 120mmminus1

120582 (A) (MoK120572) 071073119863

1199092168Mgmminus3



2) = 0134





N

O

ONi

HN

S

NN

N

OO

N N

HN

S

Ni

(2) (3)

NH2 NH2




2S2donor set





3 The 1H NMR




3)




(a) (b)












2 50 0110 50 27 times 10minus3

3 50 0105 52 29 times 10minus3










50value of



50



















0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time


DPPHBHTVitC


250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)


119886minus 120576


with complex 2




580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


N

O

ONi

HN

S

NN

N

OO

N N

HN

S

Ni

(2) (3)

NH2 NH2




2S2donor set





3 The 1H NMR




3)




(a) (b)












2 50 0110 50 27 times 10minus3

3 50 0105 52 29 times 10minus3










50value of



50



















0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time


DPPHBHTVitC


250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)


119886minus 120576


with complex 2




580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


(a) (b)












2 50 0110 50 27 times 10minus3

3 50 0105 52 29 times 10minus3










50value of



50



















0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time


DPPHBHTVitC


250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)


119886minus 120576


with complex 2




580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of

















0 10 20 30 40 50 6000

05

10

15

20

25

Abso

rban

ce at

517

nm

Time


DPPHBHTVitC


250 260 270 280 290

10

15

20

(b)

(a)

000000 000002 000004 000006

000204060810

Abso

rban

ce

Wavelength (nm)

Δ120576 a

fΔ120576 b

f

[DNA](120583M)


119886minus 120576


with complex 2




580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


580 600 620 640 6600

10

20

30

40

50

60

70Ab

sorb

ance

Wavelength (nm)

(a)

580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(b)

560 580 600 620 640 660 6800

10

20

30

40

50

60

Abso

rban

ce

Wavelength (nm)

(c)


0 10 20 30 40 50 6004

05

06

07

08

09

10

[complex][DNA]

II 0







Complexes a119870

119887Mminus1 b


119898















2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of








2O2or the complexes


4 Conclusions



Nicked DNASC DNA O2

(LB)

MPAH2O2

NiIIII(L)2(LB) DNA

DNA

OH∙


Lane 3 6 2 1 4 5 7 8 9

NC

SC



Acknowledgments


References



3COO)(H

2O)]



























































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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