Research Article Ferrocene-Based Bioactive Bimetallic...

Research ArticleFerrocene-Based Bioactive Bimetallic Thiourea ComplexesSynthesis and Spectroscopic Studies

Shafqat Ali12 Ghulam Yasin1 Zareen Zuhra1 Zhanpeng Wu1 Ian S Butler2

Amin Badshah3 and Imtiaz ud Din3

1Key Laboratory of Carbon Fiber and Functional Polymers Beijing University of Chemical Technology Ministry of EducationBeijing 100029 China2Department of Chemistry McGill University Montreal QC Canada H3A 2K63Department of Chemistry Quaid-i-Azam University Islamabad 45320 Pakistan

Correspondence should be addressed to Zhanpeng Wu drzhanpengwubuctgmailcom

Received 29 July 2015 Revised 17 September 2015 Accepted 17 September 2015

Academic Editor Zhe-Sheng Chen

Copyright copy 2015 Shafqat Ali 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

Bioactive 111015840-(441015840-di-ferrocenyl)di-phenyl thiourea and various metal complexes of this ligand have been successfully synthesizedand characterized by using physicoanalytical techniques such as FT-IR and multinuclear (1H and 13C) NMR spectroscopy alongwith melting point and elemental analyses The interaction of the synthesized compounds with DNA has been investigated byusing cyclic voltammetric and viscometric measurements The intercalation of the complexes into the double helix structure ofDNA is presumably occurring Viscosity measurements of the complexes have shown that there is a change in length and this isregarded as the least ambiguous and the most critical test of the binding model in solution The relative potential of the complexesas anti-bacterial antifungal and inhibition agents against the enzyme alkaline phosphatase EC 3131 has also been assessed andthe complexes were found to be active inhibitors

1 Introduction

One of the essential goals in medical community is tointroduce the new anticancer and antimicrobial therapeuticagents Cancer treatment using metal-based drugs is one ofthe very effective strategies for example platinum drugs cis-platin carboplatin and oxaliplatin are routinely used in theclinic to kill cancerous cells but their use has also been limiteddue to inherent and acquired resistance and the presenceof a number of dose-limiting side effects [1 2] The searchfor better metal-based drugs having the ability to overcomeproblems of drug resistance and side effects associated withplatinum based chemotherapy constitutes the foundation ofbioorganometallic chemistry Ferrocene and its derivativeshave played an important role as potential chemotherapeuticsin particular considerable attention has been paid to theirantitumour anti-inflammatory antimicrobial cytotoxic andDNA cleaving agents with respect to cancer cells [1ndash4] Thestability electroactivity and high spectroscopic activity of

ferrocene-based organometallics make them promising can-didates for many biological applications [5] The presence ofthe ferrocenyl moiety enhances activity due to its reversibleredox behaviour and increases cell permeability due to itslipophilic nature [6] It has been reported that when ferrocenewas incorporated into tamoxifen the anticancer activity ofthe drug is enhanced [7] Ferrocene derivatives may bind totheDNAvia both covalent andnoncovalentmodes of interac-tion The anticancer activity of ferrocene derivatives is foundto be dependent on the oxidation state of iron in the ferrocenemoiety with some results indicating that the Fe(II) ferrocenylcompound is more active than Fe(III) ones [8] The resultsof the study on ferrocifen as one of the Fe(II) compoundsindicate that the ferrocifens act by changing the conformationof the receptor protein [9] Binding of ferrocifen to ER120573 isthought to lead to its dimerization followed by attachment ofthe dimerized species to a particular region ofDNAThe elec-tron transfer reaction involving the ferrocenium ion in vivoor the ferrocifen-ER120573 complex may generate reactive oxygen

Hindawi Publishing CorporationBioinorganic Chemistry and ApplicationsVolume 2015 Article ID 386587 9 pageshttpdxdoiorg1011552015386587

2 Bioinorganic Chemistry and Applications

species (ROS) such as hydroxyl radicals (∙OH) ROS pro-duced can cause damage toDNA [10] andmay also be respon-sible for anticancer activity through the formation of radicalmetabolites that bring about biological damage in the cancercell [11 12]

Many researchers have reported that thiourea-basedcomplexes show effective results against various biologicalactivities due to the presence of thiocarbonyl moiety whichaffect biochemical action by the lipophilicityhydrophilicityand electronic properties of the compounds [13ndash16] Hereinwe report the synthesis characterization and investigationof DNA interaction enzymatic studies and antibacterial andantifungal activities of various metal complexes of ferrocene-based thiourea andwe believe that this studywill provide use-ful information on various biological domains and thus willbe very helpful to the design of new drug

2 Materials and Methods

21 Chemicals and Instrumentation Ferrocene hydrochlo-ric acid sodium nitrite acetonitrile dimethyl sulfoxide(DMSO) ethanol diethyl ether carbon disulfide triethy-lamine metal salts (Pd Ag Cd Zn Hg etc) ammoniumformate zinc-dust and alkaline phosphatase (ALP EC 3131)were obtained from E Merck and Aldrich (Pakistan) Allsolvents were dried and purified before use according to thereportedmethods [18] Elemental analyses (CHNS) were per-formed using an in-house instrument Leco CHNS-932 Ele-mental Analyzer Melting points were measured using a BIOCOTE Model SMP10 melting point apparatus The FT-IRspectra (4000ndash400) cmminus1 were obtained using KBr disks ona Thermo Scientific Nicolet-6700 FT-IR spectrometer TheNMR spectra of the complexes were recorded using a BrukerAvance 300MHz NMR spectrometer

22 Synthesis of 111015840-(441015840-Di-ferrocenyl)di-phenyl Thiourea(Ft) 3-Ferrocenylanilinewas synthesized in accordancewiththe methodology reported earlier [17] An ethanolic solutionof 3-ferrocenylaniline (20mmol) was added dropwise toa solution of carbon disulfide (10mmol) containing a fewdrops of triethylamine in an ice bath (0ndash5∘C) and thenreaction mixture was stirred overnight at room tempera-ture The progress of the reaction was monitored by TLCAfter completion the reaction mixture was filtered off andthe residue was recrystallized from acetonitrile to obtainthe symmetrical ferrocene-based thiourea ligand (Ft) Yield65 mp 180∘C Molecular formula (Mol wt) is found asC33H24Fe2N2S (596) FT-IR (] cmminus1) Fe-Cp (490 cmminus1) NH

(3354 cmminus1) sp2 CH(3084 cmminus1) C=CAr (1587 cmminus1)meta-disubstituted benzene (883 cmminus1) C=S (740 cmminus1) 1H NMR(300MHz CDCl

3) 120575 409 (s 5H C

5H5) 436 (s 2H C

5H4)

466 (s 2H C5H4) 759 (s 1H C

6H4) 722 (d 1H 119869 = 78Hz

C6H4) 742 (t 1H 119869 = 78Hz C

6H4) 734 (t 1H 119869 = 78Hz

C6H4) 799 (s 1H NH) ppm13C NMR (75MHz CDCl

3) 120575 6975 6935 6667 8399

13715 12278 14154 12472 12953 12233 17977 ppm Ele-mental analysis Cal() C 6646 H 473 N 470 S 538Found () C 6639 H 474 N 468 S 534

23 Synthesis of Metal Complexes The target compounds (1ndash5) were synthesized by the following general procedure

An acetonitrile solution of ferrocene-based thiourea (Ft)was added dropwise to an acetonitrile solution of the appro-priate metal salt (Zn Cd Hg Pd and Ag) in a 1 2 moleratio and the reaction mixture was stirred for 4ndash6 h at roomtemperature the extent of the reaction was monitored byTLC After completion of the reaction the mixture wasfiltered off and the residue was isolated This solid materialwas dissolved in dichloromethane and then recrystallizedusing 119899-hexane chloromethanemixture (1 3)Unfortunatelythe crystals obtained were not of sufficient quality for single-crystal X-ray diffraction analysis

24 FT-IR and Multinuclear (1H and 13C) NMR Studies TheFT-IR and multinuclear (1H and 13C) NMR spectral data forthe complexes are as follows

For compound 1 with molecular formula (Mol wt)found as C

66H56Cl2Fe4N4S2Zn (1328) FT-IR (] cmminus1) Fe-

Cp (486 cmminus1) NH (3204 cmminus1) sp2 CH (2962 cmminus1) C=CAr (1525 cmminus1) meta-disubstituted benzene (880 cmminus1) C=S(724 cmminus1) 1H NMR (300MHz DMSO) 120575 406 (s 10HC5H5) 434 (s 4H C

5H4) 473 (s 4H C

5H4) 771 (s 2H

C6H4) 727 (d 2H 119869 = 75Hz C

6H4) 733 (t 2H 119869 =

75Hz C6H4) 736 (d 2H 119869 = 75Hz C

6H4) 981 (s 2H

NH) ppm 13CNMR (75MHz DMSO) 120575 6985 6935 66568518 13985 12194 13988 12270 12882 12173 17718 ppmElemental analysis Cal() C 5965 H 425 N 422 S 483Found () C 5971 H 421 N 419 S 483 Yield 45 andmp 240∘C

For compound 2 with molecular formula (Mol wt)C66H56Cl2Fe4N4S2Cd (1376) FT-IR (] cmminus1) Fe-Cp

(484 cmminus1) NH (3290 cmminus1) sp2 CH (3097 cmminus1) C=C Ar(1590 cmminus1) meta-disubstituted benzene (885 cmminus1) C=S(728 cmminus1) 1H NMR (300MHz DMSO) 120575 409 (s 10HC5H5) 435 (s 4H C

5H4) 466 (s 4H C

5H4) 760 (s 2H

C6H4) 722 (d 2H 119869 = 75Hz C

6H4) 734 (t 2H 119869 = 78Hz

C6H4) 741 (d 2H 119869 = 75Hz C

6H4) 1008 (s 2H NH) ppm

13C NMR (75MHz DMSO) 120575 6985 6850 6575 830513620 12172 14051 12126 12859 12365 17556 ppmElemental analysis Cal() C 5761 H 410 N 407 S 466Found () C 5744 H 414 N 404 S 465 Yield 60 andmp 230∘C

For compound 3 with molecular formula (Mol wt) cal-culated as C

68H56Cl2Fe4HgN6S2(1464) FT-IR (] cmminus1) Fe-

Cp (490 cmminus1) NH (3091 cmminus1) sp2 CH (2928 cmminus1) C=CAr (1576 cmminus1) meta-disubstituted benzene (893 cmminus1) CN(2353 cmminus1) C=S (631 cmminus1) 1HNMR (300MHz DMSO) 120575422 (s 10H C

5H5) 429 (s 4H C

5H4) 462 (s 4H C

5H4)

677 (s 2H C6H4) 644 (d 2H 119869 = 72Hz C

6H4) 711 (t

2H 119869 = 78Hz C6H4) 721 (d 2H 119869 = 75Hz C

6H4) 1016

(s 2HNH) ppm 13CNMR (75MHz DMSO) 120575 6885 68606585 8315 13631 12185 14062 12139 12860 12376 1766514462 ppm Elemental analysis Cal() C 5414 H 386 N583 S 438 Found () C 5411 H 389N 583 S 442 Yield70 and mp 240∘C

For compound 4 with molecular formula (Mol wt)found asC

66H56Cl2Fe4N4PdS2(1370) FT-IR (] cmminus1) Fe-Cp

Bioinorganic Chemistry and Applications 3


5H4) 473 (s 4H C

5H4) 771 (s 2H

C6H4) 725 (d 2H 119869 = 78Hz C

6H4) 732 (t 2H 119869 =

69Hz C6H4) 739 (d 2H 119869 = 69Hz C

6H4) 981 (s 2H


For compound 5 with molecular formula (Mol wt)C66H56Fe4N5O3S2Ag (1361) FT-IR (] cmminus1) Fe-Cp

(483 cmminus1) NH (3290 cmminus1) sp2 CH (3083 cmminus1) C=C Ar(1583 cmminus1) meta-disubstituted benzene (881 cmminus1) NO-asym (1580 cmminus1) NO-sym (1541 cmminus1) C=S (724 cmminus1) 1HNMR (300MHz DMSO) 120575 403 (s 10H C

5H5) 436 (s 4H

C5H4) 477 (s 4H C

5H4) 749 (s 2H C

6H4) 716 (d 2H

119869 = 69Hz C6H4) 734 (t 2H 119869 = 75Hz C

6H4) 742 (d 2H

119869 = 75HzC6H4) 1022 (s 2HNH) ppm 13CNMR (75MHz

DMSO) 120575 6989 6928 6691 8473 13898 12382 140441227 12953 11638 17243 ppm Elemental analysis Cal()C 5818 H 411 N 514 S 471 Found () C 5819 H413 N 519 S 474 Yield 50 and mp 230∘C

25 DNA Binding Studies by Cyclic Voltammetry and Vis-cometry Cyclic voltammetric (CV) measurements were per-formed in a single compartment cell with a three-electrodeconfiguration using an Eco Chemie Auto lab PGSTAT 12potentiostatgalvanostat (Utrecht The Netherlands) instru-ment equipped with the electrochemical software packageGPES 49 The three-electrode system consisted of referenceelectrode RE-1B silver-silver chloride (AgAgCl) saturatedwith sodium chloride (NaCl) of length 70mm and outerdiameter of 60mm (ALS category number 012167) a Beck-man platinum wire of thickness 05mmwith an exposed endof 10mm as the counter electrode and a bare glassy carbonelectrode (surface area of 0071 cm2) as working electrodeThe voltammogram of a known volume of the test solutionwas recorded in the absence of calf thymus DNA (CT-DNA)after flushing out oxygen by purging with argon gas for10min just prior to each experiment The procedure wasthen repeated for systems with constant concentration of thecompounds Ft and 1ndash5 (1mM) and increasing concentrationof CT-DNA (1mL of 20 40 120583M) All the sample solutionswere prepared in 20 aqueous DMSO and buffered at pH 7by phosphate buffer (01MNaH

2PO4+ 01MNaOH) 01mM

potassium chloride (KCl) was used as supporting electrolyteThe working electrode was cleaned after every electrochem-ical assay [19] The stock solution of CT-DNA (200 120583M)was prepared by using doubly distilled water and stored at4∘C The concentration of CT-DNA was determined by UVabsorbance at 260 nm (molar coefficient L of CT-DNA wastaken as 6600Mminus1 cmminus1) The nucleotide to protein (NP)ratio of 185 was obtained from the ratio of absorbance at260 and 280 nm (A260A280 = 185) providing evidence forprotein-free DNA [20]

Viscosity measurements were carried out using OswaldViscometer maintained at a constant temperature at 250 plusmn01∘C in a thermostatic bath A series of solutions weremade with varying concentration of DNA and constantconcentration of the compound Flow times were measuredwith a digital stopwatch and each solution of the complexeswas measured three times and an average flow time was cal-culated Data are presented as 120578120578

0versus binding ratio [com-

pound][DNA] where 120578 is the viscosity of DNA in the pres-ence of complex and 120578

0is the viscosity of DNA alone All the

experimentswere conducted in 01Mphosphate buffer (pH7)at 25∘C and the results were the average of three experimentalmeasurements

26 Enzyme Inhibition Studies The basic principle of thisstudy is that the alkaline phosphatase in the sample catalyzesthe hydrolysis of colorless p-nitrophenyl phosphate (p-NPP)to give p-nitrophenol and inorganic phosphate At the pHof the assay (alkaline) the p-nitrophenol is in the yellowphenoxide form The rate of absorbance increase at 405 nmis directly proportional to the alkaline phosphatase activity inthe sample Synthesized compoundsFt and 1ndash5were screenedfor their inhibitory activity against the enzyme alkaline phos-phatase EC 3131 The enzyme activity was monitored spec-trophotometrically at constant temperature (25∘C) throughthe increase in absorbance at 405 nmwhich is associatedwiththe hydrolysis of the substrate para-nitrophenyl phosphate(pNPP) The reaction was started by addition of 40120583L ofthe enzyme to 2mL of an assay system in DMSO containing2mM pNPP in 005M Na

2CO3-NaHCO

3buffer (pH 100) at

different concentrations of the complexes Absorption mea-surements were recorded using a Beckman U-2020 spec-trophotometer

27 Antibacterial Assay Thesuccessful locally isolated patho-gens from (1) urinary tract infections (indigenous uropath-ogens) that is Klebsiella pneumonia and Escherichia coli and(2) other hospital acquired infections that is Staphylococcusaureus and Micrococcus luteus were examined for antibac-terial activities All synthesized compounds were tested by areported method with minor modifications (agar well diffu-sion assay) [17 21] where imipenem was used as the standardantibiotic [22] The whole experiment was performed at pH7 using appropriate concentration of reagents andMcFarlandsolution as turbidity standard Using a micropipette 30 120583Lof each compound was poured in their respective wells Theincubated time was 24 h at 37∘C The zone of inhibition ()was calculated for each compound and compared with thestandard antibiotic

28 Antifungal Assay Antifungal screening of the synthe-sized compounds (Ft and 1ndash5) was carried out againstAspergillus niger Terbinafine was used as standard drug [23]Different concentrations of each compound (3mgmL 5mgmL and 20mgmL) were prepared in 100mL of DMSOTubes were loaded with solutions of each compound stan-dard drug (negative) and positive control (DMSO) in thegrowth medium by using a micropipette Fungal spores weretransferred to each growth culture test tube during assay


FeFe

EtOH

Fe HNS

NHFe

Metal saltacetonitrile

FeHN

S

NHFe

FeNH

S

NHFe

MCl

Cl

M = Zn(II) Cd(II) Hg(II) Pd(II) Ag(I)

(Ft)

CS2

NH2NO2

Et2O + PTC

Et3N

+HCl(aq) + NaNO2 (aq)

H2N

Scheme 1 Synthesis of ferrocene-based bimetallic thiourea complexes

with maintaining pH 4 [24] These tubes were incubated athuman body temperature (37∘C) for one week The sameprocedure was repeated for 3 times to get better and meanresults and it was found that most significant results wereobtained for concentration of 20mgmL After required timeof incubation the zones of inhibition were measured and thepercentage of fungal inhibition was calculated and comparedwith the standard drug

3 Results and Discussion

111015840-(441015840-Di-ferrocenyl)di-phenyl thiourea was synthesizedby the reaction of 3-ferrocenylaniline and carbon disulfide inthe presence of triethylamine as a base Complexes 1ndash5 weresynthesized bymixing the thiourea ligand and differentmetalsalts in a 1 2 mole ratio (Scheme 1) Compounds Ft and 1ndash5are quite stable in moist air The molecular structures of thesynthesized compounds were established on the basis of dataobtained by elemental analysis and spectroscopic studies likemultinuclear (1H and 13C) NMR and FT-IR

31 Spectroscopic Studies

311 NMR Spectroscopy Representative 1H NMR data forthe compounds are given in the experimental section Themarker peak for Ft is the N-H signal that is shifted from370 (3-ferrocenylaniline) to 799 proving the formation ofthe symmetrical ferrocene-based thiourea A downfield shiftin N-H resonance was observed between C-N bonds Theunsubstituted C

5H5ring of ferrocene appears as a singlet in

the 1H NMR spectrum at 120575 410 whereas the ortho- and

metaprotons on the substituted Cp ring are present at 120575 465and 120575 433 respectively which split into three peaks onformation of the compound One singlet for the five protonsof one Cp ring is at 120575 409 ppm and there are two pseudotriplets at 120575 433 and 120575 465 ppm with 119869-values of 62HzThis splitting of one peak into three peaks provides evidencefor the attachment of the substituent of the one Cp ring ofthe ferrocene For complexes 1ndash5 the N-H signal of the Ftbecame less intense upon coordination and it is shifted down-field from the position in the free ligand The deshielding isrelated to an increase of 120587-electron density in the C-N bondupon coordination and it may be due to the development ofhydrogen bonding between the H of N-H and the Cl of themetal The appearance of the N-H signal shows that ligand iscoordinated to the metals via sulfur of the Ft ligand A smalldifference in chemical shift is observed in other hydrogenatoms due to 120587-character All the protons in the complexescan be identified and the total number of protons estimatedfrom the peak heights of the integration curves agrees wellwith the expected molecular formulae

The 13C NMR spectral data are also presented in exper-imental section The C=S peak appeared at 17779 ppm andall other peaks within the range confirm the synthesis of FtFor complexes 1ndash5 the 120575 (C=S) resonance of the ligand in thecomplex is shifted upfield by about 2 ppm as compared to thefree ligand The upfield shift is attributed to the lowering ofthe 120575 (C=S) bond strength producing a partial double bondcharacter in the C-N bond The shift difference of the C=Sresonance may be related to the strength of the metal-sulfurbond A small deshielding effect is observed for the othercarbon atoms due to an increase in the 120587-character of theC-N bond


312 Infrared Spectroscopy Important IR data for the com-pounds are presented in experimental section The charac-teristic bands were observed ] (C=S) at 740 cmminus1 ] (N-H)for the secondary amine in this case at 3354 cmminus1 ] (metadisubst benzene) at 883 cmminus1 ] (C-H) aromatic at 3084 cmminus1] (C=C) aromatic at 1587 cmminus1 and ] (Fe-Cp) at 490 cmminus1These bands indicate the formation of Ft The shift of thebands from those for the initial compound confirms the prod-uct formation For complexes 1ndash5 characteristic bands wereexpected in ranges indicated ] (C=S) around 729ndash750 cmminus1N-H 3204ndash3220 cmminus1 and Fc-Cp near 478ndash486 cmminus1 Thereare low frequency shifts in the ] (C=S) and ] (N-H)bands when compared to those of the free ligand

32 DNA Binding Studies through Cyclic Voltammetry Inves-tigations of drug-DNA interactions have great importance inlife science [25] Interest in understanding the association ofdrug molecules with duplex DNA has been developed in thehope of understanding the mode of binding [26] The non-covalent interactions of a drug with DNA may involve threepossible modes of interaction intercalation groove bindingand electrostatic interactions [27] There are different tech-niques which can be used to demonstrate the mode of inter-action and the DNA binding parameters One of the mostsophisticated and sensitive techniques is cyclic voltammetryVoltammetric measurements were performed in a singlecompartment cell with a three-electrode configuration withthe objective of understanding the redox behavior and theDNA binding affinities of Ft 1 and 4 [28ndash30] The mea-surements were carried out with increasing concentration ofcalf thymus DNA (1mL of 20 120583M 40 120583M) against constantconcentration (1mM) of Ft 1 and 4The voltammogramwasrecorded in the absence and presence of CT-DNA in samplesolutions On addition of increasing concentration of CT-DNA into a 1mM solution of Ft 1 and 4 a drop in current119894pa and a shift in anodic potential are observed (as shown inFigures 1 2 and 3)

The shift in peak potential is used to investigate modeof interaction between Ft 1 and 4 and DNA The slightlypositive shift in the peak potential is indicative intercalationof the compounds into double helical structure of DNA Thebinding ratio of reduced and oxidized species is calculatedaccording to the following equation [19 31]

Eb∘ minus Ef ∘ = 005916 log(119870red119870oxd) (1)

where Eb∘ and Ef ∘ are the formal potentials of the free andbound forms of drug respectivelyThe positive shift indicatesintercalation with DNA for Ft 1 and 4 The drop in currentis attributed to diffusion of the drug into the double helicalDNA resulting in the formation of a supramolecular complexAs the supramolecular complex is formed the number ofelectrons transferred is decreased and hence the drop off incurrent The increase in molecular weight of the compound(due to adduct formation with DNA) also justifies the ideathat heavy molecules migrate slowly to the electrode and so

c

a

025 03 035 04 045 05 055 0602E (V) versus SCE

minus0000005

minus0000003

minus0000001

0000001

0000003

0000005

0000007

i(A

)

Figure 1 Cyclic voltammograms of 1mM compound Ft recordedat 100mVs potential sweep rate on glassy carbon electrode at 25∘Cin the absence (a) and presence of 1mL of 20120583M 40120583M withincreasing concentration of CT-DNA (b-c) in 20 aqueous DMSObuffer at pH 70 supporting electrolyte 01M KCl

a

c

035 04 045 05 055 0603E (V) versus SCE

minus0000007

minus0000002

0000003

0000008

0000013

i(A

)

Figure 2 Cyclic voltammograms of 1mM compound 1 recorded at100mVs potential sweep rate on glassy carbon electrode at 25∘Cin the absence (a) and presence of 1mL of 20120583M 40120583M withincreasing concentration of CT-DNA (b-c) in 20 aqueous DMSObuffer at pH 70 supporting electrolyte 01M KCl

a decrease in current is observed The binding constant isdetermined using the following equation [32]

1

[DNA]=119870 (1 minus 119860)

1 minus (119894119894119900)minus 119870 (2)

where119870 is the binding constant 119894 and 119894119900are the peak currents

with and without CT-DNA and 119860 is the proportionalityconstant The plot of 1[DNA] versus 1(1 minus 119894119894

119900) yields

binding constants and is listed in Table 1The DNA binding affinity of 3-ferrocenylaniline has

already been reported by our research group [17] The DNAbinding affinity of 3-ferrocenylaniline is greater than that ofFt 1 or 4 This difference may be attributed to the mixtureof binding modes that is the ferrocenyl moiety binds elec-trostatically to the negatively charged phosphate of the DNA


a

c

minus55E minus 06

minus35E minus 06

minus15E minus 06

00000005

00000025

00000045

00000065

00000085

00000105

i(A

)

03 04 05 0602E (V) versus SCE

Figure 3 Cyclic voltammograms of 1mM compound 4 recordedat 100mVs potential sweep rate on glassy carbon electrode at 25∘Cin the absence (a) and presence of 1mL of 20 120583M 40 120583M withincreasing concentration of CT-DNA (b-c) in 20 aqueous DMSObuffer at pH 70 supporting electrolyte 01M KCl

Table 1 Binding constant and binding energy values of 1ndash4 and 3-ferrocenylaniline [17]

Compound Binding constant (Mminus1) minusΔ119866 (kJmol)Ft 343 times 103 19411 463 times 103 20232 483 times 103 20413 457 times 103 20354 585 times 103 20873-Ferrocenylaniline 939 times 103 2167

backbone and there is intercalation of the planar phenylmoiety into the base pair pockets The binding constants of1ndash4 and 3-ferrocenylaniline are listed in Table 1

The free binding energy is calculated from the equationminusΔ119866 = RT ln119870 The negative value of free binding energyof 1ndash4 and 3-ferrocenylaniline in kJmol at 25∘C shows thespontaneity of compound-DNA interaction [17] as listed inTable 1 while compound 5 showed almost same behavior ascompound 3

33 DNABinding Studies throughViscometry Another usefultechnique to prove intercalation is the viscositymeasurementwhich is sensitive to the length change of DNA due tothe lengthening of DNA helix as the base pair pocketsare widened to accommodate the binding molecule Thistechnique is regarded as the least ambiguous and the mostcritical test of the binding mode in solution under appro-priate conditions (constant temperature at 250 plusmn 01∘Cin a thermostatic bath) The plots reveal negative changesin (120578120578

0) with increasing concentration of all compounds

The graph between relative specific viscosity (1205781205780) and

[compound][DNA] for 1 and 5 is shown as representationin Figures 4 and 5 This mode of action is suggestive of

01 02 03 04 050[Compound][DNA]

11005

1011015

1021025

1031035

104

120578120578

0

Figure 4 Effect of increasing concentration of compound 1 on therelative viscosity of DNA at 25∘C [DNA] = 30 120583M and [compound-1] = 5ndash25 120583M

005 01 015 02 025 03 035 040[Compound][DNA]

1

1005

101

1015

102

1025

103

1035

120578120578

0


intercalation that may cause lengthening of the DNA chain[33]

34 In Vitro Inhibition Studies of Alkaline Phosphatase Theeffect of various concentrations of compounds Ft and 1ndash5(10 120583L 20120583L 40 120583L and 60 120583L) on the activity of the enzymealkaline phosphatase EC 3131 was studied for the hydrolysisof p-nitrophenyl phosphate (pNPP) Alkaline phosphatasecatalyzes the transfer of phosphate groups to water (hydroly-sis) or alcohol (transphosphorylation) using a wide variety ofphosphomonoesters and is characterized by high pH optimaand a broad substrate specificity [34] Here we have practicalevidence that the presence of different metals resulted in thedeactivation of the enzyme of 40 120583L as concentration Theactivity of enzyme was markedly decreased by increasingthe concentration of the compounds The activity of theenzyme (alkaline phosphatase) is presented in Figure 6

35 Antibacterial Activity In vitro evaluation of antibacterialactivity was successfully carried out The experiments wererepeated three times and the results are reported as means ofat least three determinations and the results are summarizedin Table 2 As evident from Table 2 Ft and 1ndash5 exhibited


Table 2 Antibacterial activity of Ft and 1ndash5

Chemical codesStaphylococcus aureus Klebsiella pneumoniae Micrococcus luteus Escherichia coli

(2) (119866 +ve) (1) (119866 minusve) (2) (119866 +ve) (1) (119866 minusve)Radius (mm) value Radius (mm) value Radius (mm) value Radius (mm) value

Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17

Table 3 Antifungal activity of Ft and 1ndash5

Compound codes Concentration(mg100mL)

Negative controlgrowthDMSO (cm)

Culture length (incontrol) (cm)

Fungal growth length (insample)

inhibition of fungalgrowth

Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200

Std drugs = Terbinafine (100)

Alkaline phosphatase

Activ

ity o

f enz

yme (

)

0102030405060708090

100

Ft 1 2 3 4 5BlankCompound codes

Blank

20120583L10120583L 60120583L

40120583L

Figure 6 Enzymatic studies (alkaline phosphatase) of compoundsFt and 1ndash5

significant inhibitory activity against the two strains Staphy-lococcus aureus and Micrococcus luteus as compared tostandard drug (imipenem) at the tested concentration

36 Antifungal Activity Table 3 summarizes the antifungalactivity of the compounds against pathogenic yeast speciesThe results reveal that all the compounds had promising anti-fungal activities against Aspergillus niger and poor activitiesagainst other yeastsThese results suggest that the compoundhas effective activities against selective yeasts Iron is essentialfor microorganisms as a trace nutrient Moreover severalstudies had reported that iron containing organometalliccompounds showed good antimicrobial activities [35]

4 Conclusion

Ferrocene incorporated bimetallics (1ndash5) have been synthe-sized and successfully characterized During DNA binding


studies the shift in formal potential reveals themode of inter-action between the complexes and DNA Compounds Ft1 and 4 undergo intercalation into the double helix structureof DNA and this result is also supported by viscometricmeasurements These complexes have been checked for theiralkaline phosphatase activity in the presence and absence ofinhibitor which shows that by the addition of inhibitor theactivity of enzyme decreases and at higher concentration it iscompletely inhibited Compounds Ft and 1ndash5 are biologicallyactive against Gram-positive bacteria (S aureus and Mluteus) Gram-negative bacteria (E coli and K pneumoniae)and selective yeast A niger

Conflict of Interests

The authors declare that there is no conflict of interests

Acknowledgments

Shafqat Ali acknowledges the Department of Microbiol-ogy Quaid-i-Azam University Pakistan and Department ofChemistry McGill University Montreal QC Canada fortheir support

References

[1] D Savage N Neary G Malone S R Alley J F Gallagher andP TM Kenny ldquoThe synthesis and structural characterization ofnovel N-meta-ferrocenyl benzoyl amino acid estersrdquo InorganicChemistry Communications vol 8 no 5 pp 429ndash432 2005

[2] Z PetrovskiMR PNorton deMatos S S Braga et al ldquoSynthe-sis characterization and antitumor activity of 12-disubstitutedferrocenes and cyclodextrin inclusion complexesrdquo Journal ofOrganometallic Chemistry vol 693 no 4 pp 675ndash684 2008

[3] H Parveen F Hayat A Salahuddin and A Azam ldquoSynthesischaracterization and biological evaluation of novel 6-ferro-cenyl-4-aryl-2-substituted pyrimidine derivativesrdquo EuropeanJournal of Medicinal Chemistry vol 45 no 8 pp 3497ndash35032010

[4] R H Fish and G Jaouen ldquoBioorganometallic chemistry struc-tural diversity of organometallic complexes with bioligandsand molecular recognition studies of several supramolecularhosts with biomolecules alkali-metal ions and organometallicpharmaceuticalsrdquoOrganometallics vol 22 no 11 pp 2166ndash21772003

[5] D Savage G Malone J F Gallagher Y Ida and P T MKenny ldquoSynthesis and structural characterization of N-para-ferrocenyl benzoyl amino acid ethyl esters and the X-ray crystalstructures of the glycyl and (plusmn)-2-aminobutyric acid derivativeFc C6H4CONHCH(C

2H5)CO2Etrdquo Journal of Organometallic

Chemistry vol 690 no 2 pp 383ndash393 2005[6] A Mooney A J Corry D OrsquoSullivan D K Rai and P T

M Kenny ldquoThe synthesis structural characterization and invitro anti-cancer activity of novelN-(3-ferrocenyl-2-naphthoyl)dipeptide ethyl esters and novel N-(6-ferrocenyl-2-naphthoyl)dipeptide ethyl estersrdquo Journal of Organometallic Chemistry vol694 no 6 pp 886ndash894 2009

[7] A J Corry A Goel S R Alley et al ldquoN-ortho-ferrocenyl ben-zoyl dipeptide esters synthesis structural characterization and

in vitro anti-cancer activity of N-ortho-(ferrocenyl)benzoyl-glycine-L-alanine ethyl ester andN-ortho-(ferrocenyl)benzoyl-L-alanine-glycine ethyl esterrdquo Journal of Organometallic Chem-istry vol 692 no 6 pp 1405ndash1410 2007

[8] M F R Fouda M M Abd-Elzaher R A Abdelsamaia and AA Labib ldquoOn the medicinal chemistry of ferrocenerdquo AppliedOrganometallic Chemistry vol 21 no 8 pp 613ndash625 2007

[9] Y-F Yuan L-Y Zhang J-P Cheng and J-T Wang ldquoElec-trochemical behaviour of ferrocenyl-containing acyl thioureaderivativesrdquo Transition Metal Chemistry vol 22 no 3 pp 281ndash283 1997

[10] J-Z Liu B-A Song H-T Fan et al ldquoSynthesis and in vitrostudy of pseudo-peptide thioureas containing 120572-aminophos-phonate moiety as potential antitumor agentsrdquo European Jour-nal of Medicinal Chemistry vol 45 no 11 pp 5108ndash5112 2010

[11] Z Zhong R Xing S Liu L Wang S Cai and P Li ldquoSynthesisof acyl thiourea derivatives of chitosan and their antimicrobialactivities in vitrordquo Carbohydrate Research vol 343 no 3 pp566ndash570 2008

[12] N Khan B Lal A Badshah et al ldquoDNA binding studies of newferrocene based bimetallicsrdquo Journal of the Chemical Society ofPakistan vol 35 no 3 pp 916ndash921 2013

[13] S Hussain A Badshah B Lal et al ldquoNew supramolecularferrocene incorporatedNN1015840-disubstituted thioureas synthesischaracterization DNA binding and antioxidant studiesrdquo Jour-nal of Coordination Chemistry vol 67 no 12 pp 2148ndash21592014

[14] V G Vaidyanathan and B U Nair ldquoSynthesis characterizationand binding studies of chromium(III) complex containing anintercalating ligand with DNArdquo Journal of Inorganic Biochem-istry vol 95 no 4 pp 334ndash342 2003

[15] S Ali A A Altaf A Badshah et al ldquoDNA interaction Antibac-terial and Antifungal studies of 3-nitrophenylferrocenerdquo Jour-nal of the Chemical Society of Pakistan vol 35 no 3 pp 922ndash928 2013

[16] X-B Chen Q Ye Q Wu Y-M Song R-G Xiong and X-Z You ldquoThe first organometallic carbonyl tungsten complex ofantibacterial drug norfloxacinrdquo Inorganic Chemistry Communi-cations vol 7 no 12 pp 1302ndash1305 2004

[17] S Ali A Badshah A A Ataf Imtiaz-ud-Din B Lal andK M Khan ldquoSynthesis of 3-ferrocenylaniline DNA interac-tion antibacterial and antifungal activityrdquoMedicinal ChemistryResearch vol 22 no 7 pp 3154ndash3159 2013

[18] D B G Williams and M Lawton ldquoDrying of organic solventsquantitative evaluation of the efficiency of several desiccantsrdquoJournal of Organic Chemistry vol 75 no 24 pp 8351ndash83542010

[19] C-L Bian Q-X Zeng L-J Yang H-Y Xiong X-H Zhangand S-F Wang ldquoVoltammetric studies of the interaction ofrutin with DNA and its analytical applications on theMWNTsndashCOOHFe

3O4modified electroderdquo Sensors and Actuators B

Chemical vol 156 no 2 pp 615ndash620 2011[20] A Shah M Zaheer R Qureshi Z Akhter and M Faizan

Nazar ldquoVoltammetric and spectroscopic investigations of 4-nitrophenylferrocene interacting with DNArdquo SpectrochimicaActa Part A Molecular and Biomolecular Spectroscopy vol 75no 3 pp 1082ndash1087 2010

[21] M Sonmez I Berber and E Akbas ldquoSynthesis antibacterialand antifungal activity of some new pyridazinone metal com-plexesrdquo European Journal of Medicinal Chemistry vol 41 no 1pp 101ndash105 2006


[22] A Saeed U Shaheen A Hameed and S Z H Naqvi ldquoSynthe-sis characterization and antimicrobial activity of some new 1-(fluorobenzoyl)-3-(fluorophenyl)thioureasrdquo Journal of FluorineChemistry vol 130 no 11 pp 1028ndash1034 2009

[23] F Shaheen A Badshah M Gielen et al ldquoIn vitro assessmentof cytotoxicity anti-inflammatory antifungal properties andcrystal structures of metallacyclic palladium(II) complexesrdquoJournal of Organometallic Chemistry vol 695 no 3 pp 315ndash3222010

[24] M Nath Sulaxna X Song G Eng and A Kumar ldquoSyn-thesis and spectral studies of organotin(IV) 4-amino-3-alkyl-124-triazole-5-thionates in vitro antimicrobial activityrdquo Spec-trochimica ActamdashPart A Molecular and Biomolecular Spec-troscopy vol 70 no 4 pp 766ndash774 2008

[25] H Ilkhani M R Ganjali M Arvand and P Norouzi ldquoElectro-chemical and spectroscopic study of samarium ion interactionwith DNA in different pHsrdquo International Journal of Electro-chemical Science vol 5 no 2 pp 168ndash176 2010

[26] Y-J Liu and C-H Zeng ldquoSynthesis and DNA interactionstudies of ruthenium(II) complexes with isatino[1 2-b]-1489-tetraazatriphenylene as an intercalative ligandrdquo TransitionMetal Chemistry vol 34 no 4 pp 455ndash462 2009

[27] B S P Reddy S M Sondhi and J W Lown ldquoSynthetic DNAminor groove-binding drugsrdquo Pharmacology amp Therapeuticsvol 84 no 1 pp 1ndash111 1999

[28] A Shah A M Khan R Qureshi F L Ansari M F Nazar andS S Shah ldquoRedox behavior of anticancer chalcone on a glassycarbon electrode and evaluation of its interaction parameterswith DNArdquo International Journal of Molecular Sciences vol 9no 8 pp 1424ndash1434 2008

[29] S Shujha A Shah Zia-ur-Rehman et al ldquoDiorganotin(IV)derivatives of ONO tridentate Schiff base synthesis crystalstructure in vitro antimicrobial anti-leishmanial and DNAbinding studiesrdquo European Journal of Medicinal Chemistry vol45 no 7 pp 2902ndash2911 2010

[30] Zia-ur-Rehman A Shah N Muhammad S Ali R Qureshiand I S Butler ldquoSynthesis characterization and DNA bindingstudies of penta- and hexa-coordinated diorganotin(IV) 4-(4-nitrophenyl)piperazine-1-carbodithioatesrdquo Journal of Organo-metallic Chemistry vol 694 no 13 pp 1998ndash2004 2009

[31] M S Ibrahim ldquoVoltammetric studies of the interaction ofnogalamycin antitumor drug with DNArdquo Analytica ChimicaActa vol 443 no 1 pp 63ndash72 2001

[32] N Raman K Pothiraj and T Baskaran ldquoDNA interactionantimicrobial electrochemical and spectroscopic studies ofmetal(II) complexes with tridentate heterocyclic Schiff basederived from 21015840-methylacetoacetaniliderdquo Journal of MolecularStructure vol 1000 no 1ndash3 pp 135ndash144 2011

[33] M S Ibrahim I S Shehatta and A A Al-Nayeli ldquoVoltammet-ric studies of the interaction of lumazine with cyclodextrins andDNArdquo Journal of Pharmaceutical and Biomedical Analysis vol28 no 2 pp 217ndash225 2002

[34] J G Zalatan T D Fenn and D Herschlag ldquoComparative enzy-mology in the alkaline phosphatase superfamily to determinethe catalytic role of an active-site metal ionrdquo Journal of Molecu-lar Biology vol 384 no 5 pp 1174ndash1189 2008

[35] A Imtiaz-ud-Din M Mazhar K M Khan M F Mahon andKCMolloy ldquoStudies of bimetallic carboxylates their synthesischaracterization biological activity and X-ray structurerdquo Jour-nal of Organometallic Chemistry vol 689 no 5 pp 899ndash9082004

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


species (ROS) such as hydroxyl radicals (∙OH) ROS pro-duced can cause damage toDNA [10] andmay also be respon-sible for anticancer activity through the formation of radicalmetabolites that bring about biological damage in the cancercell [11 12]

Many researchers have reported that thiourea-basedcomplexes show effective results against various biologicalactivities due to the presence of thiocarbonyl moiety whichaffect biochemical action by the lipophilicityhydrophilicityand electronic properties of the compounds [13ndash16] Hereinwe report the synthesis characterization and investigationof DNA interaction enzymatic studies and antibacterial andantifungal activities of various metal complexes of ferrocene-based thiourea andwe believe that this studywill provide use-ful information on various biological domains and thus willbe very helpful to the design of new drug

2 Materials and Methods

21 Chemicals and Instrumentation Ferrocene hydrochlo-ric acid sodium nitrite acetonitrile dimethyl sulfoxide(DMSO) ethanol diethyl ether carbon disulfide triethy-lamine metal salts (Pd Ag Cd Zn Hg etc) ammoniumformate zinc-dust and alkaline phosphatase (ALP EC 3131)were obtained from E Merck and Aldrich (Pakistan) Allsolvents were dried and purified before use according to thereportedmethods [18] Elemental analyses (CHNS) were per-formed using an in-house instrument Leco CHNS-932 Ele-mental Analyzer Melting points were measured using a BIOCOTE Model SMP10 melting point apparatus The FT-IRspectra (4000ndash400) cmminus1 were obtained using KBr disks ona Thermo Scientific Nicolet-6700 FT-IR spectrometer TheNMR spectra of the complexes were recorded using a BrukerAvance 300MHz NMR spectrometer

22 Synthesis of 111015840-(441015840-Di-ferrocenyl)di-phenyl Thiourea(Ft) 3-Ferrocenylanilinewas synthesized in accordancewiththe methodology reported earlier [17] An ethanolic solutionof 3-ferrocenylaniline (20mmol) was added dropwise toa solution of carbon disulfide (10mmol) containing a fewdrops of triethylamine in an ice bath (0ndash5∘C) and thenreaction mixture was stirred overnight at room tempera-ture The progress of the reaction was monitored by TLCAfter completion the reaction mixture was filtered off andthe residue was recrystallized from acetonitrile to obtainthe symmetrical ferrocene-based thiourea ligand (Ft) Yield65 mp 180∘C Molecular formula (Mol wt) is found asC33H24Fe2N2S (596) FT-IR (] cmminus1) Fe-Cp (490 cmminus1) NH

(3354 cmminus1) sp2 CH(3084 cmminus1) C=CAr (1587 cmminus1)meta-disubstituted benzene (883 cmminus1) C=S (740 cmminus1) 1H NMR(300MHz CDCl

3) 120575 409 (s 5H C

5H5) 436 (s 2H C

5H4)

466 (s 2H C5H4) 759 (s 1H C

6H4) 722 (d 1H 119869 = 78Hz

C6H4) 742 (t 1H 119869 = 78Hz C

6H4) 734 (t 1H 119869 = 78Hz

C6H4) 799 (s 1H NH) ppm13C NMR (75MHz CDCl

3) 120575 6975 6935 6667 8399

13715 12278 14154 12472 12953 12233 17977 ppm Ele-mental analysis Cal() C 6646 H 473 N 470 S 538Found () C 6639 H 474 N 468 S 534

23 Synthesis of Metal Complexes The target compounds (1ndash5) were synthesized by the following general procedure

An acetonitrile solution of ferrocene-based thiourea (Ft)was added dropwise to an acetonitrile solution of the appro-priate metal salt (Zn Cd Hg Pd and Ag) in a 1 2 moleratio and the reaction mixture was stirred for 4ndash6 h at roomtemperature the extent of the reaction was monitored byTLC After completion of the reaction the mixture wasfiltered off and the residue was isolated This solid materialwas dissolved in dichloromethane and then recrystallizedusing 119899-hexane chloromethanemixture (1 3)Unfortunatelythe crystals obtained were not of sufficient quality for single-crystal X-ray diffraction analysis

24 FT-IR and Multinuclear (1H and 13C) NMR Studies TheFT-IR and multinuclear (1H and 13C) NMR spectral data forthe complexes are as follows

For compound 1 with molecular formula (Mol wt)found as C

66H56Cl2Fe4N4S2Zn (1328) FT-IR (] cmminus1) Fe-

Cp (486 cmminus1) NH (3204 cmminus1) sp2 CH (2962 cmminus1) C=CAr (1525 cmminus1) meta-disubstituted benzene (880 cmminus1) C=S(724 cmminus1) 1H NMR (300MHz DMSO) 120575 406 (s 10HC5H5) 434 (s 4H C

5H4) 473 (s 4H C

5H4) 771 (s 2H

C6H4) 727 (d 2H 119869 = 75Hz C

6H4) 733 (t 2H 119869 =

75Hz C6H4) 736 (d 2H 119869 = 75Hz C

6H4) 981 (s 2H


For compound 2 with molecular formula (Mol wt)C66H56Cl2Fe4N4S2Cd (1376) FT-IR (] cmminus1) Fe-Cp


5H4) 466 (s 4H C

5H4) 760 (s 2H

C6H4) 722 (d 2H 119869 = 75Hz C

6H4) 734 (t 2H 119869 = 78Hz

C6H4) 741 (d 2H 119869 = 75Hz C

6H4) 1008 (s 2H NH) ppm

13C NMR (75MHz DMSO) 120575 6985 6850 6575 830513620 12172 14051 12126 12859 12365 17556 ppmElemental analysis Cal() C 5761 H 410 N 407 S 466Found () C 5744 H 414 N 404 S 465 Yield 60 andmp 230∘C

For compound 3 with molecular formula (Mol wt) cal-culated as C

68H56Cl2Fe4HgN6S2(1464) FT-IR (] cmminus1) Fe-

Cp (490 cmminus1) NH (3091 cmminus1) sp2 CH (2928 cmminus1) C=CAr (1576 cmminus1) meta-disubstituted benzene (893 cmminus1) CN(2353 cmminus1) C=S (631 cmminus1) 1HNMR (300MHz DMSO) 120575422 (s 10H C

5H5) 429 (s 4H C

5H4) 462 (s 4H C

5H4)

677 (s 2H C6H4) 644 (d 2H 119869 = 72Hz C

6H4) 711 (t

2H 119869 = 78Hz C6H4) 721 (d 2H 119869 = 75Hz C

6H4) 1016

(s 2HNH) ppm 13CNMR (75MHz DMSO) 120575 6885 68606585 8315 13631 12185 14062 12139 12860 12376 1766514462 ppm Elemental analysis Cal() C 5414 H 386 N583 S 438 Found () C 5411 H 389N 583 S 442 Yield70 and mp 240∘C

For compound 4 with molecular formula (Mol wt)found asC

66H56Cl2Fe4N4PdS2(1370) FT-IR (] cmminus1) Fe-Cp



5H4) 473 (s 4H C

5H4) 771 (s 2H

C6H4) 725 (d 2H 119869 = 78Hz C

6H4) 732 (t 2H 119869 =

69Hz C6H4) 739 (d 2H 119869 = 69Hz C

6H4) 981 (s 2H




5H5) 436 (s 4H

C5H4) 477 (s 4H C

5H4) 749 (s 2H C

6H4) 716 (d 2H

119869 = 69Hz C6H4) 734 (t 2H 119869 = 75Hz C

6H4) 742 (d 2H




2PO4+ 01MNaOH) 01mM








2CO3-NaHCO

3buffer (pH 100) at





FeFe

EtOH

Fe HNS

NHFe


FeHN

S

NHFe

FeNH

S

NHFe

MCl

Cl


(Ft)

CS2

NH2NO2

Et2O + PTC

Et3N


H2N

















c

a

025 03 035 04 045 05 055 0602E (V) versus SCE

minus0000005

minus0000003

minus0000001

0000001

0000003

0000005

0000007

i(A

)


a

c

035 04 045 05 055 0603E (V) versus SCE

minus0000007

minus0000002

0000003

0000008

0000013

i(A

)



1

[DNA]=119870 (1 minus 119860)

1 minus (119894119894119900)minus 119870 (2)



119900) yields




a

c

minus55E minus 06

minus35E minus 06

minus15E minus 06

00000005

00000025

00000045

00000065

00000085

00000105

i(A

)

03 04 05 0602E (V) versus SCE










01 02 03 04 050[Compound][DNA]

11005

1011015

1021025

1031035

104

120578120578

0


005 01 015 02 025 03 035 040[Compound][DNA]

1

1005

101

1015

102

1025

103

1035

120578120578

0









Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17







Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200



Activ

ity o

f enz

yme (

)

0102030405060708090

100


Blank

20120583L10120583L 60120583L

40120583L




4 Conclusion






Acknowledgments


References



















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of



5H4) 473 (s 4H C

5H4) 771 (s 2H

C6H4) 725 (d 2H 119869 = 78Hz C

6H4) 732 (t 2H 119869 =

69Hz C6H4) 739 (d 2H 119869 = 69Hz C

6H4) 981 (s 2H




5H5) 436 (s 4H

C5H4) 477 (s 4H C

5H4) 749 (s 2H C

6H4) 716 (d 2H

119869 = 69Hz C6H4) 734 (t 2H 119869 = 75Hz C

6H4) 742 (d 2H




2PO4+ 01MNaOH) 01mM








2CO3-NaHCO

3buffer (pH 100) at





FeFe

EtOH

Fe HNS

NHFe


FeHN

S

NHFe

FeNH

S

NHFe

MCl

Cl


(Ft)

CS2

NH2NO2

Et2O + PTC

Et3N


H2N

















c

a

025 03 035 04 045 05 055 0602E (V) versus SCE

minus0000005

minus0000003

minus0000001

0000001

0000003

0000005

0000007

i(A

)


a

c

035 04 045 05 055 0603E (V) versus SCE

minus0000007

minus0000002

0000003

0000008

0000013

i(A

)



1

[DNA]=119870 (1 minus 119860)

1 minus (119894119894119900)minus 119870 (2)



119900) yields




a

c

minus55E minus 06

minus35E minus 06

minus15E minus 06

00000005

00000025

00000045

00000065

00000085

00000105

i(A

)

03 04 05 0602E (V) versus SCE










01 02 03 04 050[Compound][DNA]

11005

1011015

1021025

1031035

104

120578120578

0


005 01 015 02 025 03 035 040[Compound][DNA]

1

1005

101

1015

102

1025

103

1035

120578120578

0









Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17







Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200



Activ

ity o

f enz

yme (

)

0102030405060708090

100


Blank

20120583L10120583L 60120583L

40120583L




4 Conclusion






Acknowledgments


References



















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


FeFe

EtOH

Fe HNS

NHFe


FeHN

S

NHFe

FeNH

S

NHFe

MCl

Cl


(Ft)

CS2

NH2NO2

Et2O + PTC

Et3N


H2N

















c

a

025 03 035 04 045 05 055 0602E (V) versus SCE

minus0000005

minus0000003

minus0000001

0000001

0000003

0000005

0000007

i(A

)


a

c

035 04 045 05 055 0603E (V) versus SCE

minus0000007

minus0000002

0000003

0000008

0000013

i(A

)



1

[DNA]=119870 (1 minus 119860)

1 minus (119894119894119900)minus 119870 (2)



119900) yields




a

c

minus55E minus 06

minus35E minus 06

minus15E minus 06

00000005

00000025

00000045

00000065

00000085

00000105

i(A

)

03 04 05 0602E (V) versus SCE










01 02 03 04 050[Compound][DNA]

11005

1011015

1021025

1031035

104

120578120578

0


005 01 015 02 025 03 035 040[Compound][DNA]

1

1005

101

1015

102

1025

103

1035

120578120578

0









Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17







Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200



Activ

ity o

f enz

yme (

)

0102030405060708090

100


Blank

20120583L10120583L 60120583L

40120583L




4 Conclusion






Acknowledgments


References



















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of







c

a

025 03 035 04 045 05 055 0602E (V) versus SCE

minus0000005

minus0000003

minus0000001

0000001

0000003

0000005

0000007

i(A

)


a

c

035 04 045 05 055 0603E (V) versus SCE

minus0000007

minus0000002

0000003

0000008

0000013

i(A

)



1

[DNA]=119870 (1 minus 119860)

1 minus (119894119894119900)minus 119870 (2)



119900) yields




a

c

minus55E minus 06

minus35E minus 06

minus15E minus 06

00000005

00000025

00000045

00000065

00000085

00000105

i(A

)

03 04 05 0602E (V) versus SCE










01 02 03 04 050[Compound][DNA]

11005

1011015

1021025

1031035

104

120578120578

0


005 01 015 02 025 03 035 040[Compound][DNA]

1

1005

101

1015

102

1025

103

1035

120578120578

0









Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17







Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200



Activ

ity o

f enz

yme (

)

0102030405060708090

100


Blank

20120583L10120583L 60120583L

40120583L




4 Conclusion






Acknowledgments


References



















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


a

c

minus55E minus 06

minus35E minus 06

minus15E minus 06

00000005

00000025

00000045

00000065

00000085

00000105

i(A

)

03 04 05 0602E (V) versus SCE










01 02 03 04 050[Compound][DNA]

11005

1011015

1021025

1031035

104

120578120578

0


005 01 015 02 025 03 035 040[Compound][DNA]

1

1005

101

1015

102

1025

103

1035

120578120578

0









Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17







Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200



Activ

ity o

f enz

yme (

)

0102030405060708090

100


Blank

20120583L10120583L 60120583L

40120583L




4 Conclusion






Acknowledgments


References



















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of





Imipenem 18 100 20 100 18 100 20 100Ft 13 72 02 11 16 89 02 111 00 00 3 17 3 17 5 282 3 17 2 11 4 22 3 173 7 39 3 17 00 00 4 224 9 50 6 33 7 39 11 615 3 17 04 20 00 00 03 17







Ft300 1040 1100 710 3550500 1000 1100 560 49002000 1000 1100 330 7000

1300 980 1050 970 760500 950 1050 860 18002000 1000 1050 700 3330

2300 1020 1150 1020 1130500 1030 1150 950 17402000 1000 1150 810 2956

3300 1100 1200 1050 1250500 1150 1200 820 31662000 1160 1200 630 4750

4 300 1070 1050 730 3048500 1040 1050 450 5714

5300 960 1000 840 1600500 900 1000 630 37002000 950 1000 280 7200



Activ

ity o

f enz

yme (

)

0102030405060708090

100


Blank

20120583L10120583L 60120583L

40120583L




4 Conclusion






Acknowledgments


References



















































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of





Acknowledgments


References



















































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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