Inorganica Chimica Acta 400 (2013) 2–6

Contents lists available at SciVerse ScienceDi rect

Inorgan ica Chimi ca Acta

journal homepage: www.elsevier .com/locate / ica

A new bifunctional tridentate NSN ligand leading to cationic tricarbonyl fac-[M(NSN)(CO)3]+ (M = Re, 99mTc) complexes

Ioakim Mylonas a, Charalampos Triantis a, Angeliki Panagiotopoulou b, George Patsis a,Catherine P. Raptopoulou c, Aris Terzis c, Vasilis Psycharis c, Dimitri Komiotis d, Maria Pelecanou b,Ioannis Pirmettis a, Minas Papadopoulos a,⇑a Institute of Nuclear and Radiological Sciences and Technology, Energy and Safety, National Centre for Scientific Research ‘‘Demokritos’’, 15310 Ag. Paraskevi, Athens, Greece b Institute of Biosciences & Applications, National Centre for Scientific Research ‘‘Demokritos’’, 15310 Ag. Paraskevi, Athens, Greece c Institute of Advanced Materials, Physicochemical Processes, Nanotechnology and Microsystems, Department of Materials Science, National Centre for Scientific Research ‘‘Demokritos’’, 15310 Ag. Paraskevi, Athens, Greece d Department of Biochemistry and Biotechnology, University of Thessaly, 26 Ploutonos Str., 41221 Larissa, Greece

a r t i c l e i n f o

Article history: Received 3 October 2012 Received in revised form 31 January 2013 Accepted 1 February 2013 Available online 10 February 2013

Keywords:RheniumTechnetiumTricarbonyl complexes Bifunctional

0020-1693/$ - see front matter � 2013 Elsevier B.V. Ahttp://dx.doi.org/10.1016/j.ica.2013.02.001

⇑ Corresponding author. Tel.: +30 210 6503909; faxE-mail address: mspap@rrp.demokritos.gr (M. Pap

a b s t r a c t

The synthesis and characterization of the new NSN tridentate bifunctional chelating agent 3-[ 2-(20-pyri-din-2-yl-eth ylsulfanyl)ethylamino] propionic acid (as its hydrochloric salt, 1) and of its corresp onding rhenium comp lex fac-[Re(NSN)(CO)3]+, 2, is described. Both compounds were characterized by elemental analysis, IR and NMR spectroscopies and X-ray crystallography. In complex 2 the coordination geometry around rhenium is distorted octahedral with the NSN atoms participating in the coordination sphere while the carboxylate group remains free. At tracer level, the analogous complex fac-[99mTc(NSN)(CO)3]+,3, was obtained in high yield by reacting ligand 1 with the fac-[99mTc(H2O)3(CO)3]+ precursor at pH 6.5. The structure of 3 was establishe d by HPLC comparison to the prototype rhenium comp lex 2. Complex 3is stable in solution as well as in the presence of strongly coordinating agents like histidine or cysteine. Our data indicate that ligand 1 can be used as a bifunctiona l NSN chelating agent in the design of poten- tial 99mTc-radiopharmaceuticals.

� 2013 Elsevier B.V. All rights reserved.

1. Introduction

The coordina tion chemistry of technetiu m (Tc) and its surrogate rhenium (Re) has been studied extensively as a result of the impor- tance of these radiometals in the developmen t of radiopharmaceu- ticals for imaging (99mTc) and/or radiotherapy (186Re, 188Re) in Nuclear Medicine. 99mTc (t1/2 = 6 h; Ec = 140 keV) is the radionu- clide of choice for imaging, due to its ideal nuclear properties, its low cost, and widespread availabili ty. Furthermore, the introduc- tion of the high-energy beta-emitters 186Re (t1/2 = 90 h; Emax = 1.07 MeV) and 188Re (t1/2 = 17 h; Emax = 2.12 MeV) in the developmen t of therapeutic radiopharm aceuticals has made coor- dination studies on technetium and rhenium even more attractive. In fact, 99mTc and 186/188Re can be considered to be a matched pair for imaging and therapy [1].

The bifunctional strategy for the developmen t of technetium and rhenium radiopharmaceut icals has become the most widely used method for producing well-defined technetium and rhenium la- beled receptor ligands capable of highly specific in vivo localization

ll rights reserved.

: +30 210 6503829. adopoulos).

in target tissues [2]. This strategy involves the developmen t of a suit- able bifunctiona l chelating agent (BFCA) for the chelation of the radionuc lide and the conjugation of the target specific moiety. An ideal BFCA is that which is able to form stable and inert 99mTc or 186/188Re complexes in high yield, at low concentr ation. The intro- duction of the air-stable fac-[M(H2O)3(CO)3]+ (M = 99mTc or Re) pre- cursor produced by the gentle reduction of M(VII) to M(I) under 1 atm of CO, established the fac-[M(CO)3]+ core as an easily accessi- ble platform towards the synthesis of new radiopharm aceuticals [3,4]. In the fac-[M(H2O)3(CO)3]+ synthon three coordination sites are occupied by CO groups in the stable facial configuration while the remaining three coordina tion sites are occupied by water mole- cules that can be easily replaced by suitable bidentate or tridentate ligands in aqueous solution. Both bidentate and tridentat e ligands give complexes of high kinetic and thermod ynamic stability, with tridentat e ligands exhibiting faster reaction rates and better stabiliz- ing the complex against trans-chelation reactions [4,5]. A series of suitable tridentat e chelating agents which may combine an amine or aromatic N, a thioether S, and a carboxylate O as donor atoms, has been applied to produce stable hexacoordin ated complexes [6].

In the present work, we describe the synthesis and characteri za- tion of the new BFCA 3-[2-(2-pyridin-2-yl-ethylsul fanyl)

I. Mylonas et al. / Inorganica Chimica Acta 400 (2013) 2–6 3

ethylamino ]propionic acid (NSN, 1) and of the correspondi ng tricar- bonyl rhenium and technetium complexes. The new ligand 1 and the cationic fac-[Re(NSN)(CO)3]+ complex, 2 were synthesized and fully characterized by X-ray crystallograph y and other spectroscopi cmethods (Scheme 1). Chemistry was successfully transferred at the 99mTc tracer level to obtain the analogou s radioactive complex 3.

2. Experimental

2.1. Materials and methods

All reagents and organic solvents used in this study were pur- chased from Aldrich and used without further purification. Sol- vents for high-performance liquid chromatograp hy (HPLC) were HPLC-grade. They were filtered through membrane filters(0.22 lm, Millipore, Milford, MA) and degassed by a helium fluxbefore and during use. [NEt 4]2[ReBr3(CO3] was prepared accordin gto published procedure [3]. For labeling with 99mTc a kit containing 5.5 mg NaBH 4, 4 mg Na 2CO3 and 10 mg Na–K tartrate was purged with CO gas prior to addition of Na 99mTcO4, as described in the lit- erature [4]. 2-(2-Pyridin-2-yl-ethylsul fanyl)ethylamine was syn- thesized following a reported method [7].

IR spectra were recorded as KBr pellets on a Perkin–Elmer 1600 FT-IR spectrophot ometer in the region 4000–500 cm �1. The NMR spectra were recorded in DMSO- d6 at 25 �C on a Bruker 500 MHz Avance DRX spectromete r using (CH3)4Si as the internal reference. Elemental analysis for C, H and N was conducted on a Perkin–El-mer 2400 automatic elemental analyzer. HPLC analysis was per- formed on a Waters 600 chromatograp hy system coupled to both a Waters 2487 Dual k Absorbance detector and a Gabi gamma detector from Raytest. Separations were achieved on a C-18 RP col- umn (10 lm, 250 � 4 mm) eluted with a binary gradient system at a 1 mL/min flow rate. Mobile phase A was methanol containing 0.1% trifluoroacetic acid, while mobile phase B was water contain- ing 0.1% trifluoroacetic acid. The elution gradient was 0–1 min 100% B (0% A), followed by a linear gradient to 70% A (30% B) in 9 min; this composition was held for another 10 min. After a col- umn wash with 95% A for 5 min, the column was re-equilibrated by applying the initial conditions (100% B) for 15 min prior to the next injection.

2.2. Synthesis of 1

To a solution of 2-(2-pyridin-2-yl-ethylsulf anyl)ethylamine(1.82 g, 10 mmol) in 15 mL ethanol, a solution of ethyl acrylate (1 g, 10 mmol) in 15 mL ethanol was slowly added at 0 �C. The solution was stirred for 3 h at 0 �C and for an additional 24 h at room temperat ure. The solvent was subsequent ly removed under reduced pressure and the residue was dissolved in dichlorometh- ane and washed with water. The organic layer was collected and evaporated to dryness. The residue was dissolved in 20 mL NaOH 1 N, stirred for 24 h and washed with dichlorometha ne. The pH of the aqueous layer was adjusted to 3 using HCl 1 N and the sol-

CH2=CHCOOEt

N SN SNH2

1

Scheme 1. Synthesis of ligand 1

vent was evaporated. The resulting white solid was recrystal lized from tetrahyd rofuran/metha nol and crystals of 1 suitable for X- ray analysis were obtained. Yield: 2.36 g (81%), tR: 9.7 min, IR (cm�1, KBr): 1713. Anal. Calc. for C12H19ClN2O2S: C, 49.56; H, 6.59; N, 9.63. Found: C, 49.31; H, 6.40; N, 9.58%. 1H and 13C NMR data are given in Table 1.

2.3. Synthesis of 2

To a stirred solution of [NEt 4]2[ReBr3(CO)3] (77 mg, 0.1 mmol)in 7 mL methano l, a solution of 1 (29.1 mg, 0.1 mmol) in 8 mL methano l and 0.1 mL NaOH 1 N (0.1 mmol) was added. The solu- tion was refluxed and the reaction progress was monitored by HPLC. After 5 h the solvent was removed under reduced pressure the residue was washed with dichlorometha ne and crystallized by slow evaporation from methanol. Yield: 56.2 mg (93%), tR:16.7 min, IR (cm�1, KBr): 2021, 1938, 1896, 1730. Anal. Calc. for C15H18BrN2O5ReS: C, 29.80; H, 3.00; N, 4.63. Found: C, 29.69; H, 2.92; N, 4.55%. 1H and 13C NMR data are given in Table 1. Crystals suitable for X-ray analysis were obtained from recrystallization from methano l/THF.

2.4. Synthesis of 3

900 lL of a solution of the fac-[99mTc(H2O)3(CO)3]+ precursor(pH 6.5) were added to a vial containing 100 lL of a 10 �3 M solu- tion of 1 in water. The vial was sealed, flushed with N2 and heated for 20 min at 80 �C. HPLC analysis demonst rated the formation of asingle complex eluting at 16.8 min. The yield of the reaction was quantitat ive (>98%). The identity of the 99mTc-complex was estab- lished by comparative HPLC studies using samples of the well char- acterized 2 as reference. The radioactivity recovery of the HPLC column after the injections was monitored and found to be quantitat ive.

2.5. X-ray crystallog raphy

Crystals of 1 and 2 suitable for X-ray analysis were mounted in air on a Crystal Logic Dual Goniometer diffractometer using graphite monochromate d Mo Ka radiation. Unit cell dimensions were deter- mined by using the angular settings of 25 automaticall y centered reflections in the range 11 < 2h < 23 � and they appear in Table 2.Intensity data were recorded using a h–2h scan. Three standard reflections monitored every 97 reflections showed less than 3% var- iation and no decay. Lorentz, polarizati on and psi-scan absorption correctio ns (for 2 only) were applied using Crystal Logic software. The structure s were solved by direct methods using SHELXS-97 [8]and refined by full-matr ix least squares techniques on F2 usingSHELXL-97 [9]. Further crystallogra phic details of 1: 2hmax = 48 �,reflections collected/unique/ used 2541/2387 [Rint = 0.0115]/238 7, 239 parameters refined, [Dq]max/[Dq]min = 0.255/ �0.248 e/Å3, [D/r]max = 0.001, R1/wR2 (for all data) = 0.0630/0.1066 . Hydrogen atoms were located by difference maps and were refined

HN

COOH

MCO

CO

CON

S

NHCOOH

fac-[M(CO)3]+

M = Re or 99mTc

2 M = Re3 M = 99mTc

1

23

45

6

7

8

9 1011

12

and of complexes 2 and 3.

Table 11H and 13C chemical shift s (ppm) for ligand 1 and complex 2 in DMSO- d6 at 25 �C. The atom numbering used is shown in Sche me 1.

1 2 1 2

H-1 8.50 9.08 C-1 148.75 156.16 H-2 7.24 7.60 C-2 121.60 125.67 H-3 7.73 8.16 C-3 136.56 141.10 H-4 7.34 7.76 C-4 123.15 127.90 H-6 3.01 3.55, 2.83 C-5 159.25 160.37 H-7 2.94 3.57, 2.42 C-6 37.02 38.72 H-8 2.83 3.27, 2.69 C-7 30.12 27.66 H-9 3.10 2.95, 2.21 C-8 26.27 35.15 H-10 3.12 3.51, 3.25 C-9 46.20 50.54 H-11 2.72 2.71 C-10 42.24 52.84 NH 9.07 5.55 C-11 30.19 33.07

C-12 171.57 172.64 C„O 194.32,

193.13, 192.18

Table 2Summary of crystal, intensity collection and refinement data.

1 2

Empirical formula C12H19ClN2O2S C15H18BrN2O5ReS Formula weight 290.80 604.48 Temperature 293 293 Wavelength Mo Ka 0.710730 Mo Ka 0.710730 Space group P21/a P21/ca (Å) 10.235(3) 13.096(7)b (Å) 9.830(3) 11.324(6)c (Å) 15.706(6) 14.283(7)a (�) 90 90 b (�) 105.10(2) 114.99(3)c (�) 90 90 V (Å3) 1525.6(9) 1919.9(17)Z 4 4Dcalcd (Mg m�3) 1.266 2.091 Absorption coefficient l (mm�1) 0.384 8.546 Reflections with I > 2r(I) 1787 2993 R1

a 0.0403 0.0527 wR2

a 0.0941 0.1441

a w = 1/[ r2(Fo2) + (aP)2 + bP] and P = (max Fo

2,0) + 2Fc2)/3, R1 = R(|Fo| � |Fc|)/

R(|Fo|) and wR2 = {R[w(Fo2 � Fc

2)2]/R[w(Fo2)2]}1/2.

Fig. 1. 1H–1H correlation (COSY) spectrum of complex 2 in DMSO- d6 at 25 �C (rangedH 5.70–2.00). The numbering of the atoms is shown in Scheme 1.

4 I. Mylonas et al. / Inorganica Chimica Acta 400 (2013) 2–6

isotropically . Further crystallograph ic details of 2: 2hmax = 49.5 �,reflections collected/unique/ used 3437/3279 [Rint = 0.0362]/327 9, 227 parameters refined, [Dq]max/[Dq]min = 3.296/ �2.659 e/Å3, [D/r]max = 0.000, R1/wR2 (for all data) = 0.0563/0. 1486. Hydrogen atoms were introduced at calculated positions as riding on bonded atoms. All non-H atoms were refined anisotropically.

2.6. Stability studies of 3

Aliquots of 100 lL of the isolated by HPLC pure complex 3 wereadded to 900 lL of a 10 �3 M histidine or cysteine solution in PBS, pH 7.4. The mixtures were incubated at 37 �C and were analyzed by HPLC after 4 h and 24 h [10].

3. Results and discussion

3.1. Synthesis

Ligand 1 has been designed as a new tridentat e bifunctional agend. It contains three donor-atom s, the nitrogen of the pyridine ring, the sulfur of the thioether, and the nitrogen of the secondary amine capable of stabilizing the fac-[M(CO)3]+ core. In addition, it carries a carboxylate group which is expected to remain free dur-

ing complexation and can thus serve as an anchoring point for tethering a biomolec ule of interest.

Ligand 1 was easily prepared in high yield (81%) by reacting 2-(20-pyridin-2-yl-eth ylsulfanyl)ethylamine with equimolar quan- tity of ethyl acrylate in ethanol (Scheme 1) and hydrolysis of the resulting ester. Reaction of 1 with [NEt 4]2[ReBr3(CO)3] in the pres- ence of NaOH in refluxing methano l led to the formation of the cat- ionic fac-[Re(NSN)(CO)3]Br complex, 2 (Scheme 1). HPLC analysis of the reaction mixture showed the formation of a single product in excellent yield.

Compound s 1 and 2 were characterized by elemental analysis, spectroscop ic methods and X-ray crystallogra phy. They are soluble in methanol and ethanol and insoluble in ether, hexane and dichlo- romethane. Both of them are stable in the solid state and in solu- tion for months as shown by HPLC and NMR.

The infrared spectra of complex 2 demonstrate strong bands at 2021, 1938 and 1896 cm �1 attributed to the C„O stretch of the fac-[Re(CO)3]+ unit [11]. Furthermor e, a strong band at 1730 cm �1

indicates the presence of free carboxylate group. The NMR assignment s of ligand 1 and complex 2 in DMSO-d 6

were based on the combined analysis of a series of 1H–1H and 1H–13C correlation spectra and are presente d in Table 1. The num- bering of the atoms is shown in Scheme 1. Upon complexation of li- gand 1, the C-1 carbon adjacent to the coordinatin g nitrogen is shifted downfield by 7.4 ppm, while the correspond ing H-1 proton is shifted downfield by 0.58 ppm. These downfield shifts are typical for the coordinatio n of the heterocyclic aromatic nitrogen [12]. In addition upon coordina tion differentiation of the previously equiv- alent protons (H-6, H-7, H-8, H-9 and H-10) of the methylene groups of the chelated NSN backbone is noted. In both 1 and 2the H-6 protons are distinguishe d from the H-7 protons based on the presence of a NOESY peak with the neighboring pyridinyl H-4 proton and the assignment is confirmed through the long-range heteronu clear coupling of H-4 and C-6. In the COSY spectrum of complex 2 (Fig. 1) cross-pea ks between the NH proton and the four different iated protons on C-9 and C-10 are present. The fact that in complex 2 the NH proton appears shifted upfield by 4.2 ppm rela- tive to its position in ligand 1, indicates that after complexation the amine group is no longer involved in intermolecular hydrogen

Fig. 2. Partially labeled plot of the cation of 1 with ellipsoids drawn at the 30% probability level. Most of the hydrogen atoms have been omitted for clarity.

Fig. 3. Partially labeled plot of the cation of 2 with ellipsoids drawn at the 30% probability level. Most of the hydrogen atoms have been omitted for clarity.

Fig. 4. Plot of the 3D network of 1 due to hydrogen bonding interactions (dashedlines). Color code: Cl, black; S, dark grey; O, medium grey (large); N, large open circles; C, small open circles; H, medium grey (small). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

I. Mylonas et al. / Inorganica Chimica Acta 400 (2013) 2–6 5

bonding, in conformity with the crystal structures of 1 and 2 pre-sented below.

16 ,8V

3.2. Description of the crystal structures of 1 and 2

The molecula r structure s of 1 and 2 are shown in Figs. 2 and 3respectively ; selected bond distances and angles for 2 are listed in Table 3.

The structure of 1 consists of organic cations due to protonati on on the secondary amine group and chloride anions. The terminal car- boxylato group remains protonated and is involved in strong inter- molecular hydrogen bonds to the pyridine nitrogen of a neighbori ng cation [O2 � � �N1 (0.5 � x, 0.5 + y, 1 � z) = 2.564 Å, H2O � � �N1 =1.543 Å, O2–H2O� � �N1 = 172.2 �], thus creating zig-zag chains extending parallel to the (�101) plane. The protonated secondary amine group is also involved in strong intermolecular hydrogen bonds to the chloride anion [N2 � � �Cl (x,y,z) = 3.068 Å, H2NA � � �Cl =2.178 Å, N2–H2NA� � �Cl = 176.7 �; N2 � � �Cl (0.5 + x, 0.5 � y, z) =3.066 Å, H2NB � � �Cl = 2.216 Å, N2–H2NB� � �Cl = 164.3 �], thus gener- ating an overall 3D network (Fig. 4).

The structure of 2 consists of complex cations and bromide an- ions. The coordinatio n geometry around rhenium is distorted octa- hedral and consists of the three facially bound CO groups, the pyridine and amine nitrogen atoms as well as the sulfur atom of the tridentate ligand. The apical positions of the distorted octahe-

Table 3Selected bond distances (Å) and angles (o) for 2.

DistancesRe–C(23) 1.915(10) Re–N(1) 2.227(7)Re–C(21) 1.921(9) Re–N(2) 2.265(7)Re–C(22) 1.928(9) Re–S(1) 2.474(2)

AnglesC(23)–Re–C(21) 90.0(4) C(22)–Re–N(2) 94.7(3)C(23)–Re–C(22) 86.3(4) N(1)–Re–N(2) 86.7(2)C(21)–Re–C(22) 88.9(4) C(23)–Re–S(1) 96.6(3)C(23)–Re–N(1) 92.3(4) C(21)–Re–S(1) 173.4(3)C(21)–Re–N(1) 91.0(3) C(22)–Re–S(1) 90.8(3)C(22)–Re–N(1) 178.7(3) N(1)–Re–S(1) 89.4(2)C(23)–Re–N(2) 176.6(3) N(2)–Re–S(1) 80.2(2)C(21)–Re–N(2) 93.2(3)

dron around Re are occupied by the pyridine nitrogen atom, N(1),and the carbonyl group C(22). Rhenium lies on the equatorial plane defined by the four remaining coordinated atoms. The fac-[Re(CO)3]+ moiety in 5 can be described as OC-6-44- A in terms of the specific annotation reported by Bandoli [13] to account for the absolute configuration about the metal ion. The Re–carbonylbond distances, 1.915(10)–1.928(9) Å, are consisten t with those found in other Re–tricarbonyl complexes [14], with the largest dis- tance correspondi ng to C(22) in the apical position. The Re–N bond distances are 2.227(7) and 2.265(7) Å for N(1) and N(2), respec- tively; the latter bond distance is longer, albeit in the equatorial plane, due to the sp3 hybridiza tion of N(2) with respect to sp2

hybridiza tion of N(1). The Re–S–C–C–C–N six-membered ring in the coordination sphere adopts the chair conformation with Re and C(6) being 0.55 and 0.75 Å out of the mean plane of the remaining four atoms. The Re–S–C–C–N five-membered ring in the coordination sphere adopts the envelope conformation with C(9) being 0.65 Å out of mean plane of the remaining four atoms. In the lattice structure of 2, the bromide anion is weakly interact- ing to the amine nitrogen [N2 � � �Br (1 � x, 1 � y, 1 � z) = 3.432 Å,H2N� � �Br = 2.544 Å, N2–H2N� � �Br = 165.2 �].

3.3. Radioche mistry

The [99mTc(NSN)(CO)3]+ complex 3 was prepared by reacting li- gand 1 with the fac-[99mTc(H2O)3(CO)3]+ precursor at 80 �C for

3020100

3

min

16 ,7

m

2

Fig. 5. Comparative HPLC chromatograms for complexes 2 and 3.

6 I. Mylonas et al. / Inorganica Chimica Acta 400 (2013) 2–6

20 min at pH 6.5. The yield of the reaction was quantitative (>98%)at ligand concentr ation of 10 �4 M. The identity of 3 was established by comparison of its HPLC retention time to that of the well-cha r- acterized analogous rhenium complex 2 by applying parallel radio- metric and photometric detection (Fig. 5).

The isolated by HPLC complex 3 was incubated in saline as well as in 1 mM histidine and 1 mM cysteine solutions at 37 �C and was found to be >95% stable in all these conditions for 24 h, with no decompositi on or trans-chelat ion being observed . Therefore, com- plex 3 is expected to be stable at the physiolog ical histidine and cysteine concentrations [15].

4. Conclusions

The novel tridentate ligand 3-[2-(20-pyridin-2-y l-ethylsulfa- nyl)-ethylamino] propionic acid, 1, was synthesized and success- fully used for the synthesis of stable cationic complexes of the general formula fac-[M(NSN)(CO)3]+, (M = Re, 99mTc). The carboxyl- ate group does not coordinate to the metal and thus can be used for the conjugat ion of a target-specific biomolec ules either pre-label- ing or post-labelin g.

Appendix A. Supplementar y material

CCDC 903474 and 903475 contain the supplem entary crystallo- graphic data for this paper. These data can be obtained free of charge from The Cambridge Crystallo graphic Data Centre via www.ccdc.cam .ac.uk/data_req uest/cif .

References

[1] (a) S.S. Jurisson, J.D. Lydon, Chem. Rev. 99 (1999) 2205; (b) J.R. Dilworth, S.J. Parrott, Chem. Soc. Rev. 27 (1998) 43.

[2] (a) S. Liu, Adv. Drug Deliv. Rev. 60 (2008) 1347; (b) S. Liu, Chem. Soc. Rev. 33 (2004) 445.

[3] R. Alberto, A. Egli, U. Abram, K. Hagetschweiler, V. Gramlich, P.A. Schubiger, J. Chem. Soc., Dalton Trans. 19 (1994) 2815.

[4] R. Alberto, R. Schibli, A. Egli, A.P. Schubiger, J. Am. Chem. Soc. 120 (1998) 7987. [5] R. Alberto, J.K. Pak, D. van Staveren, S. Mundwiler, P. Benny, Biopolymers 76

(2004) 324. [6] (a) R. Alberto, Top. Curr. Chem. 252 (2005) 1;

(b) R. Schibli, R. La Bella, R. Alberto, E. Garcia-Garayoa, K. Ortner, U. Abram, P.A. Schubiger, Bioconjugate Chem. 11 (2000) 345.

[7] A. Buschauer, F. Lachenmayr, W. Schunack, Pharmazie 47 (1992) 86. [8] G.M. Sheldrick, SHELXS-97, Structure Solving Program, University of Göttingen,

Germany, 1997. [9] G.M. Sheldrick, SHELXS-97, Crystal Structure Refinement, University of

Göttingen, Germany, 1997. [10] T. Tsotakos, C. Tsoukalas, G. Patsis, A. Panagiotopoulou, N. Nikolic ´ , D. Jankovic ´ ,

D. Djokic ´ , C.P. Raptopoulou, A. Terzis, D. Papagiannopoulou, M. Pelecanou, M. Papadopoulos, I. Pirmettis, Inorg. Chim. Acta 377 (2011) 62–68.

[11] (a) S. Alves, A. Paulo, J.D.G. Correia, A. Domingos, I. Santos, J. Chem. Soc., Dalton (2002) 4714; (b) A. Vitor, S. Alves, J.D.G. Correia, A. Paulo, I. Santos, J. Organomet. Chem. 689 (2004) 4764.

[12] (a) O. Karagiorgou, G. Patsis, M. Pelecanou, C.P. Raptopoulou, A. Terzis, T. Siatra-Papastaikoudi, R. Alberto, I. Pirmettis, M. Papadopoulos, Inorg. Chem. 44 (2005) 4118; D. Papagiannopoulou, C. Tsoukalas, G. Makris, C.P. Raptopoulou, V. Psycharis, L. Leondiadis, E. Gniazdowska, Inorg. Chim. Acta 378 (2011) 333.

[13] G. Bandoli, F. Tisato, A. Dolmella, S. Agostini, Coord. Chem. Rev. 250 (2006)561.

[14] L. Wei, J. Babich, W.C. Eckelman, J. Zubieta, Inorg. Chem. 44 (2005) 2198–2209.and refs. therein.

[15] (a) M.D. Armstrong, U. Stave, Metabolism 22 (4) (1973) 561; (b) J.P. Milsom, M.Y. Morgan, S. Sherlock, Metabolism 28 (4) (1979) 313.