Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 115 (2013) 641–647

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Spectrochimica Acta Part A: Molecular andBiomolecular Spectroscopy

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Some new [(thione)2Au(diamine)]Cl3 complexes: Synthesis,spectroscopic characterization, computational and in vitrocytotoxic studies

1386-1425/$ - see front matter � 2013 Elsevier B.V. All rights reserved.http://dx.doi.org/10.1016/j.saa.2013.06.086

⇑ Corresponding author.E-mail address: aisab@kfupm.edu.sa (A.A. Isab).

Bassem A. Al-Maythalony a, M. Monim-ul-Mehboob a, Muhammad Altaf a, Mohammed I.M. Wazeer a,Anvarhusein A. Isab a,⇑, Saleh Altuwaijri b, Ayesha Ahmed c, Vikram Dhuna d, Gaurav Bhatia d,Kshitija Dhuna e, Sukhdev Singh Kamboj e

a Department of Chemistry, King Fahd University of Petroleum & Minerals, Dhahran 31261, Saudi Arabiab Clinical Research Laboratory, SAAD Research Development Center, SAAD Specialist Hospital, Al-Khobar 31952, Saudi Arabiac Anatomic Pathology, University of Dammam &, King Fahd Hospital of the University, Saudi Arabiad Department of Biotechnology, DAV College, Amritsar 143 001, Punjab, Indiae Department of Molecular Biology and Biochemistry, Guru Nanak Dev University, Amritsar 143 005, Punjab, India

h i g h l i g h t s

� Gold(III) complexes of the type[(thione)2Au(diamine)]Cl3

synthesized.� Cytotoxicity of complexes measured

on C6 glioma cell lines.� Spectroscopic data are evaluated by

comparisons with calculated data.

g r a p h i c a l a b s t r a c t

New Au(III) complexes of the type [(thione)2Au(diamine)]Cl3 are reported, where thione = 1,3-imidazoli-dine-2-thione (Imt), 1,3-Diazinane-2-thione (Diaz) and diamine = diaminoethane (en), 1,3-diaminopro-pane (pn) or 1,4-diaminobutane (bn). The solid state IR as well as 13C and 15N NMR data indicate thatAu(III) center is bonded via sulfur of thiocarbonyl S@C� site of the thiones and also chelated by the dia-mines from the trans side of coordinated thiones.

[(Imt)2Au(en)3+ complex

a r t i c l e i n f o

Article history:Received 15 February 2013Received in revised form 28 April 2013Accepted 24 June 2013Available online 3 July 2013

Keywords:Au(III) thione complexes13C and 15N solid state NMRC6 glioma cell lines

a b s t r a c t

Recent advances in oncology are focused on developing new complexes of gold(III) with various ligandsthat show augmented anti-proliferative potential and reduced toxicity as compared to cis-platin. In thisstudy, new Au(III) complexes of the type [(thione)2Au(diamine)]Cl3 are reported, where thione =1,3-imidazolidine-2-thione (Imt), 1,3-Diazinane-2-thione (Diaz) and diamine = 1,2-diaminoethane (en),1,3-diaminopropane (pn) or 1,4-diaminobutane (bn). The solid state IR as well as 13C and 15N NMR dataindicate that Au(III) center is bonded via sulfur of thiocarbonyl S@C� site of the thiones and also chelatedby the diamines from the trans side of coordinated thiones. Spectroscopic data are evaluated by compar-isons with calculated data from the built and optimized structure by GAUSSIAN 09 at the RB3LYP levelwith LanL2DZ bases set. These new Au(III) complexes based on mixed thione and diamine ligands arevery similar to the square planar structure of tetracoordinate [Au(en)2]Cl3complex. In this study, cytotox-

642 B.A. Al-Maythalony et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 115 (2013) 641–647

icity data for these gold(III) complexes against C6 glioma cell lines are also reported, and the results indi-cate some complexes have cytotoxicity comparable to cis-platin.

� 2013 Elsevier B.V. All rights reserved.

Introduction

Gold complexes such as aurothioglucose (Solganol), aurothiom-alate (Myocrisin) and triethylphosphine-gold(I)tetraacetylthioglu-cose (Auranofln), have a long history of their successful use inthe treatment of rheumatoid arthritis [1]. In recent years, muchinterest has been focused to investigate anti-proliferative potentialof these and related gold compounds. Many such examples haveshown promising in vitro and in vivo results [2,3]. Furthermore,gold complexes have demonstrated a strong inhibition of the en-zyme thioredoxinreductase (TrxRa), which is involved in tumorcell proliferation [4].

Gold–thiones have found great interest in recent research dueto potent gold sulfur interaction which found several medicinalapplications [5,6]. Thione heterocyclic rings with various metalions, have received much interest [7–9]. Myocrisin and Solganolused for the treatment of rheumatoid arthritis are gold–sulfurcompounds [10,11]. Considerable work has been carried out on thi-one complexes with gold(I) [12], nonetheless, very little is knownabout Au(III) complexes with thiones.

Heterocyclic thiones as ligands in metal complexes have at-tracted considerable attention because of their relevance to biolog-ical systems and the versatility in their coordination modes. Thesethiones are potentially ambidentate or multi-functional donorswith either the exocyclic S or heterocyclic N atom available forcoordination, thus yielding a variety of interesting complexes withgeometries of variable nuclearities and great structural diversity[13].

Parish et al. [14] tested a gold(III) complex [AuCl2(damp)]in vitro against a range of microbial strains and several human tu-mor lines, where it displays differential cytotoxicity similar to thatof cis-platin. Against the ZR-75-1 human tumor xenograft, both[AuCl2(damp)] and cis-platin showed limited activity. Wong et al.[15] synthesized a new class of mono- and binuclearcyclometalat-edgold(III) complexes bearing the trans dicarbanion of 2,6-diphen-ylpyridine. The molecular structures of [Au(CNC)(Spy-2)](Spy-2 = 2-mercaptopyridine),[Au2(CNC)2(l-dppm)](ClO4)2, [Au2(-CNC)2(l-dppe)](ClO4)2, and [Au(CNC)PPh3]ClO4, were determinedby X-ray crystallography.

Milovanovic et al. [16] have evaluated in vitro the cytotoxicproperties of [Au(en)Cl2]+ against chronic lymphocytic leukemiacells and shown comparable cytotoxicity profiles to cis-platin.Casini et al. [17] have studied the anticancer activities in a seriesof [Au(en)2]Cl3 type complexes. They reported that [Au(en)2]Cl3

showed similar activities compared to cis-platin.Volarevic et al. [18] investigated the cytotoxic activity of newly

synthesized gold(III) complexes [AuCl(2)(en)]+, [AuCl2(SMC)]+,[AuCl2(DMSO)2]+(en: ethylenediamine, SMC: S-methyl- L-cysteineand DMSO: for dimethylsulfoxide) in 4T1 mouse breast cancer cellline in vitro using MTT colorimetric technique and in vivo usingexternal caliper. Among the tested gold(III) complexes,[Au(en)Cl2]+ showed best cytotoxic effects in vitro. Magheriniet al. [19] have shown that a group of structurally diverse goldcompounds are highly cytotoxic toward a panel of 36 human tu-mor cell lines through a variety of biochemical mechanisms.

Arsenijevic et al. [20] tested the cytotoxicity of gold(III) com-plexes i.e. [Au(en)Cl2]+ [Au(dach)Cl2]+ and [Au(bipy)Cl2]+ on A549human lung carcinoma epithelial cell line using cytotoxic andapoptotic assays. The results showed that all tested gold(III)complexes displayed cytotoxic effect on A549 cells. Among the

tested gold(III) complexes, [Au(bipy)Cl2]+ showed the best cyto-toxic effects. Langner et al. [21] determined the cytotoxicity ofthe stable water-soluble carboxylato gold(I) [(Ph3P)AuO2C(CH2-

OCH2)nH], (n = 1,2,3), to compare the intrinsic antineoplasticactivity of gold(I) towards the HeLa (human cervix epithelioid)cancer cell line ATCC CCL-2, resting lymphocytes and phytohae-magglutinin (PHA)-stimulated lymphocytes. Gold(I) carboxylatocomplexes were also 2–5 times more toxic against PHA-stimulatedlymphocyte cultures than to HeLa cancer cell. Gold(I) complexeswere almost as cytotoxic as [Au(PPh2CH2CH2PPh2)2]Cl andcis-platin.

Serratrice et al. [22] showed important cytotoxic effects of thenew gold(III) complexes:[Au{2-(20-pyridyl)imidazolate}Cl2]and[Au{2,6-bis(20-benzimidazolate)pyridine}(OCOCH3)] and the mono- and binuclear gold(I) complexes: [Au{2-(20-pyridyl)imidaz-ole}(PPh3)]PF6, [Au(2-phenylimidazolate)(DAPTA)] (DAPTA = 3,7 -diacetyl-1,3,7-triaza-5-phosphabicyclo[3.3.1] nonane), [(PPh3Au)2

(2-R-imidazolate)]PF6 (R = 2-C5H4N, Ph). Interestingly, the com-pounds containing the 2-(20-pyridyl)imidazolate ligand showedselectivity towards cancer cells with respect to the non-tumori-genic ones, with the dinuclear compound [(PPh3Au)2(2-R-imidazo-late)]PF6 being the most active. Sivaram et al. [23] reported thecytotoxic activities of a series of Au(I) and Au(III) mono-, hom-obis-, and heterobis(carbene) complexes in vitro with theNCI-H1666 non-small cell lung cancer cell line. The cationicbis(carbene) complexes show generally better cytotoxicity in com-parison to cis-platin.

We have recently reported that the [Au(en)Cl2]Cl complexcauses minimal histological changes in kidney and liver of rats,reflecting its relative safety as compared to other clinically estab-lished anti-neoplastic drugs [24]. Likewise, our research group re-ported [Au(en)2]Cl3 as a promising candidate as an anti-canceragent [25]. For that reason, [Au(en)2]Cl3 might be a promising che-mo-preventative and chemo-therapeutic agent against human gas-tric carcinogenesis.

Such optimistic literature reports on Au(III) complexes triggerus to carry out the synthesis of [Au(diamine)Cl2]+ with thiones suchas Imt and Diaz. Accordingly, we have synthesized mixed ligandgold(III) complexes by using different diamines and thiones. Subse-quently, these synthesized complexes have been characterized byCHNS analysis, solid IR as well as solid and solution NMR spectro-scopic methods. Finally, Au(III) thione complexes have also beenevaluated as cytotoxic agents against C6 glioma cell lines. A gliomais kind of cancer that originates in the brain or spine. The mostcommon site of gliomas is the brain. Gliomas compose about 30%of all brain and central nervous system (CNS) cancers and 80% ofall malignant brain cancers [26].

Experimental

Synthesis of Au(III)thione complexes

Firstly, [(diamine)AuCl2]Cl complexes were prepared accordingto the literature [27]. Later, [(diamine)Au(thione)2]Cl were synthe-sized as follows: 0.6 mmol of [(diamine)AuCl2]Cl was dissolved in20 mL distilled H2O (solution 1). 1.2 mmol of thione was dissolvedin 40 mL methanol (solution 2). Both solutions were mixed inround bottom flask and refluxed at 60 �C for 4 h. The resulting solu-tion was filtered, concentrated and allowed to precipitate. The re-sulted precipitate is dried and filtrate is kept in refrigerator for

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crystallization. The yield of the complexes was obtained in therange of 51–66%. These synthesized complexes using the ligandsgiven in Scheme 1 were subsequently characterized by meltingpoint and elemental analysis as given in Table 1 while structuralanalysis for such complexes were carried out by FTIR spectroscopyand solid-state as well as solution NMR .

Spectroscopic characterization of Au(III)thione complexes

The solid state FTIR spectra of the ligands and their complexeswere recorded on a Perkin Elmer FTIR 180 spectrophotometerusing KBr pellets over the range 4000–400 cm�1 results are givenin Table 2.

The 1H NMR spectra were obtained on a Jeol JNM-LA 500 NMRspectrometer operating at a frequency of 500 MHz. The 13C NMRspectra were obtained at the frequency of 125.65 MHz with 1Hbroadband decoupling at 297 K. The spectral conditions were:32 k data points, 0.967 s acquisition time, 1.00 pulse delay and45� pulse angle. The 13C chemical shifts are given in Table 3. Timedependent 1H NMR spectra of 0.01 M [Au(en)(Imt)2]3+ in d6-DMSOran up to 48 h showed no change in 1H NMR spectrum indicatingthe complex is very stable.

Natural abundance 13C solid state NMR spectra were obtainedon a JEOL LAMBDA 500 spectrometer operating at 125.65 MHz(11.74 T), at ambient temperature of 25 �C. Samples were packedinto 6 mm zirconium oxide rotors. Cross polarization and highpower decoupling were employed. Pulse delay of 7.0 s and a con-tact time of 5.0 ms were used in the CPMAS experiments. Themagic angle spinning rates were from 2000 Hz to 4000 Hz. 13Cchemical shifts were referenced to TMS by setting the high fre-quency isotropic peak of solid adamantine to 38.56 ppm. Thechemical shifts are given in Table 4.

15N spectra were also recorded on the above machine, using acontact time of 5 ms and a pulse delay of 10 s. The magic anglespinning rates were between 4000 Hz and 5000 Hz. The chemicalshift of nitrogen was initially referenced with respect to liquidNH3, by setting the 15N peak in enriched solid 15NH4Cl to40.73 ppm [28] and then converted to the standard nitromethaneby a shift of �380.0 ppm [29] for ammonia. The solid-state NMRdata are given in Table 4.

(a) Imidazolidine-2-thione (Imt)

(b) 1,3-Diazinane-2-thione (Diaz)

Scheme 1. Structures and abbreviations of thione ligands used in this study.

Computational study

The structures of compounds 1 and 2 were built and optimizedusing GAUSSIAN09 program [30] at DFT/B3LYP level with LANL2DZbasis set [31,32]. Selected bond lengths and bond angles are givenin Tables 5 and 6 respectively for complexes 1 and 2.

In vitro cytotoxicity of Au(III)thione complexes on C6 glioma cells

C6 glioma cells were seeded at 2 � 104 cells/well in 100 lLDMEM (containing 10% FBS in 96-well tissue culture plate, andincubated for 72 h at 37 �C, 5% CO2 in air and 90% relative humidityin CO2 incubator in order to evaluate the in vitro cytotoxicity of[(thione)2Au(diamine)]Cl3 complexes.

160, 80, 40, 20, 10 and 5 lg/mL sample solutions were preparedin DMEM. 100 lL of each sample solution were added to the incu-bated C6 glioma cells and further incubated for 20 h. The mediumwas discarded and 100 lL DMEM containing MTT (5 mg/mL) wasadded to the same cells and again incubated in CO2 incubator at37 �C in dark for 4 h. The dark purple crystals of formazan wereproduced in the cells at the bottom of the wells.

The medium of culture from each well was aspirated cautiouslyto prevent disruption of the cell monolayer.100 lL of DMSO wasadded in each well and mixed thoroughly to dissolve the formazancrystals, producing a purple solution. The absorbance of the 96 wellplates was taken at 570 nm with Labsystems Multiskan EX-ELISA(enzyme-linked immune-sorbent assay) reader against a reagentblank.

Results and discussion

This work reports the synthesis of some [(thione)2Au(diamine)]Cl3complexes where thione = 1,3-imidazolidine-2-thione (Imt), 1,3-Diazinane-2-thione (Diaz) and diamine = 1,2-diaminoethane (en),1,3-diaminopropane (pn) or 1,4-diaminobutane (bn).

IR characterization

IR frequencies for the prepared complexes were recorded in therange 4000–400 cm�1 using KBr pellet, and the selected IR fre-quencies are given in Table 2. The vibrational frequency for thioneligands t(C=S) shows red shift upon complexation as a result of elec-tron donation from sulfur to the metal center that promote p-backdonation to thione p� orbital, this causes weakening of the CASbond [11]. On the other hand, the NAH vibrational frequency,t(N–H), was found to shift to higher wave number (blue shift) indi-cating the strengthening of NAH bond in complex compared to thefree ligand, as a result of electron donation form sulfur [33].

Solution NMR characterization

Chemical shifts at 182.11 and 45.38 ppm correspond toS@C�(free Imt) and CACH2ANH(free Imt) respectively. The downfield chemical shift for carbon of S@C�(bound Imt) from 182.1 toabout 175.8 ppm accounts for coordination of sulfur of S@C� to-wards Au(III) center. Chemical shifts at 173.34, 41.20 and19.07 ppm correspond to S@C�(free Daz), CACH2ANH(free Diaz)and CACH2AC(free Diaz) respectively. The larger down field chem-ical shift for carbon of S@C�(bound Diaz) from 173.3 to about155.5 ppm attributes the ligation of sulfur of S@C� towards Au(III)center. The smaller downfield shifts of CACH2ANH(bound Diaz)and CACH2AC(bound Diaz) also corroborates the binding of carbonof S@C�(bound Diaz) to Au(III). The evident downfield shift ofcarbon of S@C� bound thione in [(thione)2Au(diamine)]Cl3 with re-

Table 1Elemental analysis of the prepared Au(III) thione complexes.

Complex M.P. (�C) Yield (%) Found (calculated)

%C %H %N %S

1. [(Imt)2Au(en)]Cl3 201 55 17.19(16.92) 3.73(3.55) 14.77(14.80) 11.19(11.30)2. [(Imt)2Au(pn)]Cl3 218 49 18.88(18.58) 4.05(3.81) 15.00(14.45) 11.04(11.02)3. [(Imt)2Au(bn)]Cl3 172 43 20.30(20.16) 4.20(4.06) 14.19(14.11) 9.99(10.76)4. [(Diaz)2Au(en)]Cl3 240 53 21.04(20.16) 4.28(4.06) 14.30(14.11) 9.91(10.76)5. [(Diaz)2Au(pn)]Cl3 212 49 24.43(24.59) 5.06(5.31) 15.59(15.55) 10.00(9.38)6. [(Diaz)2Au(bn)]Cl3 269 44 23.18(23.31) 4.72(4.52) 13.50(13.47) 9.95(10.28)

Table 2IR frequencies, m(cm�1) of Au(III) thione complexes.

Complex t(N–H) (cm�1) t(C=S) (cm�1)

Imt 3200 5061. [(Imt)2Au(en)]Cl3 3426 w 4952. [(Imt)2Au(pn)]Cl3 3426 s 4973. [(Imt)2Au(bn)]Cl3 3400 w 496Diaz 3166 5164. [[(Diaz)2Au(en)]Cl3 3423 w 5185. [(Diaz)2Au(pn)]Cl3 3444 m 5176. [(Diaz)2Au(bn)]Cl3 3413 sh 499

Table 313C chemical shifts (ppm) for free thione ligands and Au(III) thione complexes inDMSO-d6.

Species S@C�(Imt) CACH2ANH(Imt)

Free Imt 182.11 45.381. [(Imt)2Au(en)]Cl3 175.73 45.102. [(Imt)2Au(pn)]Cl3 175.93 45.063. [(Imt)2Au(bn)]Cl3 175.94 45.06

S@C�(Diaz) CACH2ANH(Diaz) CACH2AC(Diaz)

Free Diaz 173.34 41.20 19.074. [(Diaz)2Au(en)]Cl3 166.56 36.68 18.425. [(Diaz)2Au(pn)]Cl3 166.59 36.33 18.426. [(Diaz)2Au(bn)]Cl3 166.51 38.38 18.40

Table 4Solid state 13C and 15N NMR chemical shifts of Au(III) thione complexes.

Complex 13C 15N

1. [(Imt)2Au(en)]Cl3 165.4, 46.8 �260.1, �335.82. [(Imt)2Au(pn)]Cl3 177.0, 46.6 �258.2, �342.23. [(Imt)2Au(bn)]Cl3 176.9, 46.3 �259.6, �349.34. [(Diaz)2Au(en)]Cl3 166.7, 42.7, 19.4 �262.5, �357.65. [(Diaz)2Au(pn)]Cl3 167.3, 43.5, 19.5 �262.8, �337.56. [(Diaz)2Au(bn)]Cl3 172.5, 45.2, 27.7 �258.2, �333.5

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spect to free thiones indicates coordination of thiones through sul-fur of S@C� to Au(III) center as shown in Table 3.

1H NMR for [(thione)2Au(diamine)]Cl3 complexes showed veryintricate spectra because of the strained structures of diamineligands and non-equivalency of hydrogen environments in the

Table 5Selected bond lengths (Å) for [(thione)2Au(diamine)]Cl3 for optimized structure using B3L

[(Imt)2Au(en)]3+ [(Imt)2Au(pn)]3+ [(Imt)2Au(bn)]3+

Au–N1 2.163 2.159 2.157Au–N2 2.173 2.171 2.157Au–S1 2.473 2.483 2.493Au–S2 2.471 2.482 2.493S1–C 1.825 1.824 1.829S2–C 1.824 1.823 1.829

complexes [27]. 1H NMR data were, therefore, found to be non-descriptive for the complexes.

Solid-state NMR characterization

The solid-state NMR data for the complexes are given in Table 4.The thiocarbonyl resonances in the ligands Imt and Diaz, appear at180.6 and 175.6 ppm respectively [34]. So it can be seen that thio-carbonyl carbons are shielded by about 12 to 17 ppm relative tothe free ligand, confirming the coordination through the sulfuratom of the thione group. The high frequency 15N signal is assignedto Imt and Diaz nitrogens, whereas the low frequency 15N signal isassigned to the alkyldiamine ligands. The 15N shifts of the alkyldi-amines in the complexes show low frequency shifts of 5–17 ppm,compared to that in the free ligands. These shifts indicate an in-crease in electron density at the nitrogen atom, in agreement withliterature [35].

Computational study

Built and optimized structures were analyzed computationallyfor vibrational frequencies and NMR spectra using B3LYP/LanL2DZlevel of theory, to facilitate the spectroscopic data interpretation.Selected bond length and torsion angles of the computationallyoptimized structure are presented in Tables 5 and 6, respectively(see Scheme 2).Comparisons of calculated bond lengths show thatCAS bond of thiones became longer after complexation; this agreeswith experimental IR data that shows red shift upon complexation.Additionally, CAS bonds were longer in Diaz complexes comparedto CAS bond length in Imt complexes indication of larger backdonation in Diaz complexes and stronger bond in Diaz complexesin general, this observation agree also with the higher up field shifteffect on Diaz CAS carbon as detected by 13C NMR, which clarifythe higher back donation in Diaz complexes. After optimization,the built complexes showed square planer structure around goldatom, S–Au–N angles were ranging from 173� to 179�.

In vitro cytotoxicity of Au(III)thione complexes

The EAE (Experimental Autoimmune Encephalomyelitis) wasevaluated for its antiproliferative effect using MTT assay. The [(thi-one)2Au(diamine)]Cl3 complexes inhibited the proliferation of C6glioma cells in a concentration dependent manner. IC50 values

YP/LanL2DZ.

[(Diaz)2Au(en)]3+ [(Diaz)2Au(pn)]3+ [(Diaz)2Au(bn)]3+

2.167 2.174 2.1772.167 2.164 2.1562.474 2.481 2.4822.474 2.477 2.4801.859 1.851 1.8511.859 1.851 1.852

Table 6Selected torsion Angle (�) for [(thione)2Au(diamine)]Cl3for optimized structure using B3LYP/LanL2DZ.

[(Imt)2Au(en)]3+ [(Imt)2Au(pn)]3+ [(Imt)2Au(bn)]3+ [(Diaz)2Au(en)]3+ [(Diaz)2Au(pn)]3+ [(Diaz)2Au(bn)]3+

S1–Au–N1 93.87 89.37 86.79 93.24 87.37 86.79S2–Au–N2 91.93 87.43 86.79 93.23 90.20 86.79N1–Au–N2 81.81 90.71 94.40 81.84 90.73 94.40S1–Au–S2 92.53 92.66 92.18 91.89 91.76 88.35S1–Au–N2 174.55 176.96 176.75 173.98 176.16 176.10S2–Au–N1 173.15 176.16 176.76 173.98 176.96 177.07Au–S1–C 106.76 107.14 108.57 108.65 110.14 109.41Au–S2–C 110.77 111.74 108.57 108.66 110.14 109.47

Scheme 2. Optimized geometries of [(thione)2Au(diamine)]Cl3 Complexes (1–6) at the B3LYP/LanL2DZ level of theory using Gaussian 09, Revision A. 1.

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Table 7IC50 data in micromolar for Au(III) thione complexes.

Complex IC50 (lg/mL) IC50 (lM)

1. [(Imt)2Au(en)]Cl3 42 90.210062. [(Imt)2Au(pn)]Cl3 36 75.060993. [(Imt)2Au(bn)]Cl3 73 147.8814. [(Diaz)2Au(en)]Cl3 152 316.92425. [(Diaz)2Au(pn)]Cl3 62 125.59766. [(Diaz)2Au(bn)]Cl3 61 120.1592

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were calculated from the growth curve using MTT assay (Fig. 1). Itshowed values of inhibition 42, 36, 73, 152, 62, 61 lg/mL for [(thi-one)2Au(diamine)]Cl3 complexes 1, 2, 3, 4, 5 and 6, respectively.The IC50 values 90.2, 75.1, 147.9, 316.9, 125.6 and 120.2 in lM cor-responds to complexes 1, 2, 3, 4, 5 and 6, respectively. The IC50 datain lM for [(thione)2Au(diamine)]Cl3 complexes are given inTable 7.

It is evident from IC50 values that Imt containing Au(III) com-plexes (1–3) are relatively better cytotoxic agents than Diaz con-taining Au(III) complexes (4–6). As far as cytotoxicity against C6glomia cells is concerned, the five member thione ring (Imt) isfairly better ligand than the six member thione ring (Diaz). How-ever, the six member Au(III) chelates (2 and 5) with 1,3-Diamino-propane (pn) are comparatively better cytotoxic agents than fivemember chelate (1 and 4) with 1, 2-diaminoethane (en) as wellas seven member chelate (3 and 6) with 1,4-diaminobutane (bn).It can be concluded that structural features e.g. ring size of thioneligand (Imt and Diaz) as well as Au(III) chelate size with diamine(en, pn and bn) may affect in vitro cytotoxicity of complex againstC6 glomia cell line. In this study, the interesting features of struc-ture–activity relationship (SAR) are recognized by using differentring size of thiones (Imt and Diaz), and by different chelate sizeowing to diamine ligands (en, pn and bn), incorporation in thegold(III) complexes.

The MTT results based on phase contrast micrographs showed adecrease in cell density on treatment with different concentrationsof complexes (data not shown). Significant numbers of cells werefound to be round, detached and floating at higher concentrationsand the samples were photographed after removing dead floatingcells to improve the quality of images.

Even though the antitumor efficacy of imidazolidine-2-thioneand ethylenediamine derivatives of [(thione)2Au(diamine)]Cl3 com-plexes has independently been reported, their chemotherapeuticpotential as a combined complex has not, so far, been analyzed.

Fig. 1. MTT assay of C6 glioma cell lines treated wi

The Imidazolidine-2-thione (Imt) containing complexes 1[(Imt)2Au(en)]Cl3 and 2 [(Imt)2Au(en)]Cl3 demonstrated toxicityprofiles comparable to cis-platin [36] which has been documentedto have an overall IC50 in the range of 10.9–67.0 lM. The IC50 of dif-ferent anticancer compounds specifically against C6 glioma celllines has been reported by Yildrim et al. [37]. They tested anti-pro-liferative activity of novel platinum compounds against C6 gliomacells and demonstrated an IC50 range of 16.75–28.75 lg/ml withrespect to cis-platin. Comparison with other gold(III) compoundshas scarcely been studied.

Pivetta et al. analyzed the role of imidazolidine-2-thione as ananti-cancer agent in combination with other metal ions [38]. Theyfound a high in vitro cytotoxicity of imidazolidine-2-thione copper(II) complexes in mouse neuroblastoma N2a cells. Imidazolidineinfluences apoptosis, a mechanism of programmed cell death func-tioning to eliminate damaged, mutated or cancerous cells. Chemo-therapeutic agents cause simultaneous activation of apoptoticsignals and induction of anti-apoptotic factors, such as NF-kappaB and subsequently diminish the overall efficacy of the adminis-tered drug.

Sharma et al. [39,40] described imidazoline to modulate theNF-kappa B pathway by preventing its nuclear translocation andselectively sensitizing the cancer cells toward DNA damaging

th [(thione)2Au(diamine)]Cl3 complexes (1–6).

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agent. Pretreatment of cancer cells with the imidazoline causes asignificant increase in apoptosis resulting in augmented efficacyof camptothecin and cis-platin. As far as the future perspective isconcerned, the comparative studies are required to interpret thefunctional advantages of N-alkyl substituted thiones(Imt and Diaz)and to establish structure–activity relationship (SAR) using differ-ent ring size of thiones (Imt, Diaz and Diap) incorporation in thegold(III) complexes.

Conclusions

A new series of gold(III) complexes of general formula [(thi-one)2Au(diamine)]Cl3 complexes are reported here. The solid IRas well as NMR studies show that Au(III) is bonded via S@C� siteof the thiones. The most likely structure Au(III) thione complexeswould be square planer type complexes. Since [Au(en)2]Cl3 typecomplexes are relatively more toxic towards some cancer cell lines,the [(thione)2Au(diamine)]Cl3 complexes may be a better alternatefor [Au(en)2]Cl3 type complexes. Such kind of Au(III) thione com-plexes shows cytotoxicity similar to cis-platin in C6 glioma cells.The in vitro cytotoxic study of thione Au(III) complexes againstC6 glomia cell line also leads to conclusion that the structural fea-tures e.g. ring size of thione ligand (Imt and Diaz) as well as Au(III)chelate size with diamine (en, pn and bn) may affect in vitro cyto-toxicity of complex. Further, in vivo biological studies is needed toevaluate these new complexes.

Acknowledgements

This research was supported by the King Fahd University ofPetroleum and Minerals (KFUPM) Research Committee underProject No. IN100039.

Appendix A. Supplementary material

Supplementary data associated with this article can be found, inthe online version, at http://dx.doi.org/10.1016/j.saa.2013.06.086.

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