SYNTHESIS,CHARACTERIZATION AND BIOLOGICAL EVALUATION OF

MetalBasedDrugs Vol. 7,Nr. 5, 2000

SYNTHESIS,CHARACTERIZATION AND BIOLOGICAL EVALUATION OF

Co(II), Cu(II), Ni(II) AND Zn(II) COMPLEXES WITH CEPHRADINE Zahid H. Chohan* and Maimoon F. Jaffery

Departmentof Chemistry,IslamiaUniversity,Bahawalpur,Pakistan ABSTRACT

Some Co(II), Cu(II), Ni(II) and Zn(II) complexes ofantibacterial drug cephradine have been prepared and characterized bytheirphysical,spectral and analytical data. Cephradineacts as bidentateand the complexes have compositions, [M(L)2X2] ^where [M=Co(II), Ni(II) and Zn(II), L=cephradine and X=CI2] showing octahedral geometry, and

[M(L)2]

where [M=Cu(II), L=cephradine] showing square planar geometry. In order to evaluate the effect of metal ionsupon chelation, eephradineand its complexes have been screened for their antibacterial activity against bacterial strains, Escherichia coli, Staphylococcus aureus, and Pseudomonasaeruginosa

INTRODUCTION

Evidences supporting the introduction of metallic elements in several biological processes are rapidly accumulating

r6.

Schubert⁷ and Kirschner⁸ have investigated the antibacterial, antiviral and anticancer activities of more than 25 inorganiccompounds,which includedthe metal atom aspotentially significant part of the molecule. They suggested^8’9 thatthe transfer of metal ion from the ligand to the cancer-associated viruses was a mechanism for releasingtheanticancerdruginthe locality of thetumor. Duetothe significant nature ofthe metallic ions, theirmetal complexesarenow, being included inthe search for ideal anticancer drugs. Such significantexamples ofwhich are, palladium and platinum complexes of6-mercaptopurine that destroy¹ adenocarcinomas and similarly, the complexes of dialkyldithiophosphate which reduce some tumors

1.

Awide rangeof activitiesofeventhe simplest complexesof metallic elements has been reviewed by Rosenberg²who noticedthat the viral-induced cancers allrespondtothe treatment with even thesimplest metal-amine halide complexes. It has also been demonstrated that ehelation/complexation tend to make inactive substances/ligands active and active compounds/drugs become more active and less toxic

316.

All these evidences however, highlight the need to study and evaluate more the biological applications of metallicelementsfortherapeutic potentials.

Cephradineisafirst-generation eephalosporinclass of antibiotic

7,

activelyusedagainst Gram-positivecocci, and Gram-negative bacilli. Ithas similarstructural andantibacterialrelationshiptothat ofits closely related analogue cephalexin. Itcontains the (NH), (COOH), (NH) and (C=O) ^functional groups and its molecular model reveals that its structure is suitablefor chelation/complexation. Adetailedbiologicalevaluationofthe copper(II) and Zn(II) complexes of cephalexin has been reported

.

^{In order to} evaluate the biological comparison ofboth the analogues, we, therefore, report in this paper the preparation, characterization and biological properties ofcobalt(II), copper(II), niekel(II)andZn(II)complexesofeephradinewhich maybe an addition to reveal the significance of the strong relationship between metals, or their complexes, and bactericidalactivity.

EXPERIMENTAL MaterialandMethods

Allchemicalsand solvents used were of Analargrade. Allmetal(II) salts were used as chlorides. Sodium salt of cephradinewas obtainedby reacting equimolaramount of sodium hydroxidewithcephradine. IR spectra were recorded on aPhilips Analytical PU 9800 FTIR spectrophotometer as KBr discs. UV-Visible spectra were obtained in DMF on a Hitachi U-2000 double-beam spectrophotometer. C, H and N analyses was carried outby Butterworth Laboratories Ltd. Conductance ofthe metal complexes was determined inDMF

ona Hitachi YSI-32 model conductometer. Magneticmeasurements weremadeon solidcomplexes usingthe Gouymethod.Melting pointswererecorded on aGallenkampapparatusand are uncorrected.

Preparation ofcobalt(ll)-cephrafline Complex

An ethanol solution (20 mL) of the cobalt chloride hexahydrate (0.24 g, 0.001 M) was added to a magnetically stirred sodium salt ofcephradine (0.8 g, 0.002M) in distilled water(25 mL). Themixture was refluxed for h and cooled to room temperature. On cooling, pink precipitates were formed, filtered and washed with ethanol, acetone and ether, and dried by suction. Crystallization in aqueous ethanol (70 30) gave thedesiredmetalcomplex. All othermetalcomplexeswereformed followingthe samemethod.

AntibacterialStudies

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Provided by PubMed Central

ZH. Chohan andM.F. Jaffery Synthesis, CharacterizationandBiologicalEvaluation

of

Co(II), Cu(lI),Ni(ll) andZn(II) Complexes^withCephradine Pseudomonas aeruginosa. The paper disc diffusion method was adopted for the determination of antibacterialactivity

"r4"16.

RESULTSANDDISCUSSION

All the complexes are intensely colored and stable amorphous solids, which did not have sharp melting points anddecompose above

225C.

Thecomplexesareonly solubleinDMFandDMSOand insoluble inall othercommonorganicsolvents. Theirmeltingbehavior,solubility and crystalline naturesuggestthattheyare monomers. Molar conductance values (14-18

ohmlcmZmol "l)

^ofthe complexes inDMF solution indicated¹⁹ that allof the complexesarenon-electrolytes.

Table 1.

Cephradine

Physicaland AnalyticalDataof the

Metal(!I) Complexes

Comple

M.p

B.M.

Mol.Formula

(C) (laar)

218

232-234

4.7

228’230

1.4

231-233 3.3

234’236

Dia (1)

[Co(L)2(C1)2] C32H36CoCI2N6OsS2 [956.9]

[Cu(L)2]

C32H36CuN608S2 ^[890,7]

[Ni(L)2(CI)2] C32H36NiClN608S [95617]

[Zn(L):z(CI)2] C32H36ZnCIN608S2

^[963.4]

Yield

(%)

60 62 58 62

cale. (Found)

^%

C H N CI

40.1

3.8 8.8

7.4

(39.9)(3.5)(8.9)(7.2)

43.1 4.0 9.4

(43.4)(3.9)(9.7)

40.1 3.8 8.8

7.4 (40.5)(4.1)(8.6)(7.3)

39.8 3.7 8.7 7.3 (40,

!)(3.5)(8.9)(7.5)

Examination ofthe physical model of cephradineshows that it can exhibit either ONON, OONOor OOON donor quadridentate, ONO, OOO, NNO or NNN donor tridentate and NO, OO, or NN donor bidentate behavior. Themolecularmodelstudies furtherreveal thatin no casesthe ligandcan stereochemically behave asquadridentateortridentate.

The IR spectra ofthe complexes in comparison to the uncomplexed cephradine are listed in Table 2 with some tentative important characteristic assignments

2.

The IR spectrum of the cephradine shows some characteristicbands at3345, 3240, 1770and 1745 mainly duetothe v(NH2), v(NH),v(COOH)andv(C=O) vibrations, respectively. Themetal complexes contained all the bands from cephradineand also other bands indicativeof the coordination of the ligandwiththe metal ions. Thebanddue to

v(COO)asym

at 1770 cm

"

ⁱⁿ

the spectrum of the ligand ^{shifted to} lower frequency (10-15 cm

")

in all the metalcomplexes indicative of the complexation

2.

Anew absorptionband instead, assignedtov(COO)andv(M-O) appeared at 1565-1575 and450-455 cm^"1 which were only observedin the spectraof thecomplexes. This inturn, indicatedthat the carboxyl group is coordinated tothe metalion. Also,theband dueto

v(NH2)

^at3345cm

"

^{wasfound shifted}

to lower wave number(15-20 cm

"l)

in the spectraof its metal complexes. And a new bandat415-420 cm^-l assigned to

v(M-N)

was evolved in the spectra ofthe complexes indicating² involvement of the

v(NH)

groupvianitrogeninthe coordination. Moreover, Inthe farinfraredregionanewbandat335cm¹wasfound in the spectra of the Co(II), Ni(lI) and

Zn(II)

complexes and not observed in the spectrum of the

Cu(II)

complex assigned ^to

v(M-CI)

modes respectively. It ⁱⁿ turn, indicated that two chloride atoms are also coordinated to the metal atom. These evidences however, indicated that the Co(II) Ni(II) and Zn(II) complexes possess an octahedral geometry (Fig

1A)

and Cu(II) complex to show square planar geometry H and

lC

NMR spectra of the uncomplexed cephradine and its metal complexes have been recorded in

DMSO-d6

^{with TMS} as internal reference and are summerized in Table 2 with some tentative assignments.

The spectra ofcephradine exhibited peaks as expected

za.

Signals for the NH and COOH protons in the spectra ofuncomplexed Cephradineat 1.1 and 10.8 ppm disappearedin the complexes. This indicatedthat Cephradine is coordinated to the metal ions bydeprotonation. In the complexesthe aromatic proton signals

2425

appeareddownfielddue to increasedconjugation duringcoordination

UV-Visiblespectral bands of thecomplexes arerecordedinTable 2. Thecobalt(II)complex shows magnetic moment 4.7 B.M. at room temperature (Table. 1). These high value of the magnetic moment and the stoichiometry suggestacoordinationnumber ofsixfor the centralcobalt(II) ion and an octahedral geometry.

The electronic spectrum ofthe complex is consistent with its octahedral environment around the cobalt(II) ion(Fig.lA). ^Thespectrum displaystwo bands at20,715 and28,580cm^"1 attributed to

4Tlg 4Tlg

and

4Tlg

4 2627

T2gtransitions,respectively,^{in a}high-spin octahedralgeometry

MetalBased

Drugs

Vol. 7,Nr. 5, 2000

Table 2. IR, UVand NMR

Spec

Cephra

dine/

Complex,.

Cephra dine

3345(s, NHz),³²⁴⁰ (ms, NH), ¹⁷⁷⁰ (s, COOH), ¹⁷⁴⁵ (s, c=o)

3530

(s,

NH2),1760,

1565(s,COO),⁴⁵⁰ (m, M-N),⁴²⁰(m, M-O), ³⁵⁵ (m, M- Cl)

3535(s, NH2),1755, 1565(s, COO),455 (m, M-N),⁴²⁵(m, M-O)

3535(s, NH2),1762, 1570(s, COO),⁴⁵⁵ (m,M-N),425(m, M-O), 355 (m, M- CI)

3533(s,NH2),1760, 1565(s, CO0),⁴⁵⁰ (m,M-N),⁴²²(m, M-O), ³⁵⁵ (m, M- Cl)

Xoscopic Dataof the

Metal(II)

Complexes

(cn.a,t) ^IH-NMR

(ppm)

^(DMSO-d6)

20715, 2850

15155, 18575, 30245

16570, 27375

27555

s=sharp,ms=mediumsharp,m=medium

2.3(s,6H,CH3),^4.5(d,2H, NCH),^7.1(m,4H,Ph),^7.2(m, 4H,Ph),7.4(m,2H,Ph),8.1 (d, 2H,i-Lactam), 8.3(m,4H, hetero-Ph),^8.4(m,2H,hetero-

Ph).

2.3(s,6H,CH3),4.5(d,^2H, NCH),^7.1(m,4H,Ph),^7.2(m, 4H,Ph),^7.4(m,2H,ah),^8.1 (d, 2H,

-Lactam),

8.3(m,4H, hetero-Ph),^8.4(m,2H,hetero-

Ph).

2.3

(s,

6H,CH3),^4.5(d,2H, NCH),7.1(m,4H,Ph),7.2(m, 4H,Ph),7.4(m,2H,Ph),8.1 (d, 2H,I-Lactam),^8.3(m,4H, hetero-Ph),8.4(m,2H, hetero-

Ph).

2.3(s,6H,CH3),^4.5(d,2H, NCH),^7.1 (m,4H,Ph),^7.2(m, 4H,ah),^7.4(m,2H,Ph),^8.1 (d, 2H,i-Lactam), ^8.3(m,4H, hetero-Ph),8.4(m,2H,hetero-

Ph).

2.3(s,6H,CH3),^4.5(d,2H, NCH),7.1 (m,4H,Ph),7.2(m, 4H,Ph),7.4(m,2H,Ph),8.1 (d, 2H, I$-Lactam), 8.3(m,4H, hetero-Ph),^8.4(m,2H,hetero-

Ph).

C-NMR (DMSO-d6)

(ppm)

13(CH3),

63

(CHN), 117 (-Lactam),120, 122,124 (hetero-Ph), 148, 125, 127, 128,172(C=O), 177 (COO),²¹²(l-Lactam).

13

(CH3),

63(CHN), 117 (l-Lactam),120, 122, 124 (hetero-Ph),148, 125, 127, 128, 172(C=O), 177 (COO),212(i5-Lactam).

13(CH3),

63

(CHN), 117 (l$-Lactam), 120,122, 124 (hetero-Ph), 148, 125, 127, 128, 172(C=O), 177 (COO),212

(I5-Lactam).

13 (CH3),⁶³

(CHN),’

117 (l$-Lactam), 120, 122, 124 (hetero-Ph),148, 125, 127, 128, 172(C=O), 177 (COO),²¹²

(13-Lactam).

13 (CH3),63(CHN), 117 (-Lactam), 120, 122, 124 (hetero-Ph),148, 125, 127, 128, 172(C=O),177 (COO),212(l-Lactam).

[IA] M=Co(II),Ni(II)orZn(II); ^X=CI [1B] M=Cu(II);^X=O

Z.H.Chohanand M.. F.Jaffery Synthesis, CharacterizationandBiologicalEvaluation

of

Co(ll), Cu(ll),Ni(ll) andZn(ll) Complexes^withCephradine configuration andmay be assignedto

2Big - ^2Alg

^and

^2Big -- ^2Eg

transitions respectively

2s’29.

The strong high-energyband isassignedtometal

---

^{ligandchargetransfer. Also,}the magnetic moment value(1.4 B.M) for the Cu(II) complex is foundtobe consistent withtheproposed square planar geometry (Fig 1B) forthe Cu(II)complex.

The electronic spectra ^{of the} nickel(II) complex exhibited absorption bands at 16,570 and 27,375 cm

,

attributed to

3A2g 3Tlg

(F) and

3AEg 3Tlg

(P)transitions,respectively,in anoctahedral geometry

3’3.

The calculated values of theligand fieldparameter lie inthe range reported for anoctahedral structure.Also, the valueof the magnetic^moment(3.3 B.M.)may betaken as an additional evidenceforitsoctahedralstructure.

The diamagnetic zinc(II) complex did not show any d-d bands and its spectrum is dominated only by_ the charge transferband. This charge transfer band at27,555 cm was assigned to the transition

2Eg

^---,

2T2g,

possiblyin anoctahedralenvironment

3.

On thebasisoftheaboveobservations, it istentativelysuggested thattheCo(II), Ni(II)andZn(II)complexes show an octahedralgeometry (Fig. 1A) in which the two cephradinemolecules act as bidentate and ataxial positions two chlorides are coordinated to the metal atom. In the Cu(II) complex the two cephradine molecules respectively, acting as bidentate show a square planar geometry of the complex (Fig. 1B) by possibly accommodatingthemselvesaroundthe metal atom in such awaythat astable chelateringisformed.

Antibacterial Properties

Cephradine in comparison to its metal(II) chelates was evaluated for antibacterial activity against the standard bacterial strains, Escherichia coli(a),), Staphylococcusaureus (b) and Pseudomonasaeruginosa(c) The compounds were tested at a concentration of 30 lag/0.01 mL in DMF solution using the paper disc diffusion method reported^33’34 as earlier. Thesusceptibility zones were measured in diameter (mm) and the resultsarereproduced inTable 3. The susceptibilityzones were theclearzonesaround the discs. Cephradine and its complexes individually were found to be biologically active showing various degrees ofinhibitory effects on the growth of the tested bacterial species. The antibacterial results evidently show that the complexation improved the antibacterial activity. These observations however, suggest that the metallic elements should be seriously considered in <drug designing>. Their use could provide an easy way of improving thebactericidalactivity of many drugs/antibacterial agents.

Table3. Antibacterial

Activity

^{Data of}Cephradin

e

^andits

^{Metal(II) Complexes}

Cephradine/Complex M c r o b a S _p e c e s

a b c

Cephradine ^{++ +} ++

(1)

^{+++ ++}

+++..

(2) ^{++++ +++ +++}

(3)

^{+++ ++}

++++

(4) +++

+++ ++

a Escherichia coli, b Staphylococcusaureus,

c Pseudomonas aeruginosa. Inhibition zone diametermm(%inhibition):

+,

6-10(27-45 %);

++,

10-14(45-64%);

+++,

14-18(64-82 %);

++++,

18-22(82-100 %). Percentinhibitionvalues arerelative to inhibition zone(22 mm)with O0 % inhibition.

ACKNOWLEDGEMENT

The authorsgratefullyacknowledgethehelpof theDepartment^ofMicrobiology,Qaid-e-Azam Medical College, Bahawalpur,inperforming theantibacterial studies.

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Received- September 20, 2000 Accepted: November 2, 2000

Received in revised camera-ready format: January 16, 2001