Synthesis, structure and anti-fungal activity of dimeric Ag(I) complexes containing bis-imidazole ligands

Syn hesis, s uc u e and an i- ungal ac i i y o dime ic Ag(I) complexes con aining bis-imidazole ligands Suaad Abuskhuna a , John B iody a , Malachy McCann a,* , Michael De e eux b , Ke in Ka anagh c , Julia Ba ei a Fon echa d , Vickie McKee d a Depa men o Chemis y, S . Pa ick’s College, Na ional Uni e si y o I eland Maynoo h, Maynoo h, Co. Kilda e, I eland b Dublin Ins i u e o Technology, Ca hal B ugha S ee , Dublin, I eland c Depa men o Biology, Na ional Uni e si y o I eland Maynoo h, Maynoo h, Co. Kilda e, I eland d Depa men o Chemis y, Loughbo ough Uni e si y, Loughbo ough, Leics LE11 3TU, UK Recei ed 10 No embe 2003; accep ed 4 Feb ua y 2004 Abs ac Fi e Ag(I) complexes con aining he ligands bis(imidazol-2-yl)me hane (2-BIM) and i s de i a i es we e p epa ed and [Ag2(2- BIM)2](ClO4)2and [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH we e cha ac e ised using X- ay c ys allog aphy. In each dime he wo Ag(I) ions a e wo-coo dina e and he e a e small bu defini e a gen ophilic Ag–Ag (d10–d10) in e ac ions. All o he complexes display an i- ungal ac i i y when es ed in i o agains he ungal pa hogen Candida albicans. Ó2004 Else ie L d. All igh s ese ed. Keywo ds: Bis-imidazole; Bis(imidazol-2-yl)me hane; X- ay s uc u es; Sil e (I); A gen ophilici y; An i- ungal; Candida albicans 1. In oduc ion In ou effo s o de elop new me al complexes ha would g ea ly inhibi he g ow h o he human ungal pa hogen Candida albicans [1–4] we ha e ound ha , o da e, he mos ac i e d ug es ed in aqueous media is he Ag(I) complex [Ag(phendio)2]ClO4(phendio ¼1,10- phenan h oline-5,6-dione) [5]. As pa o hese ongoing s udies in o he syn hesis and in i o es ing o new me al- based an i- ungal d ugs we epo he e he p epa a ion o fi e Ag(I) complexes con aining bis(imidazole) ligands. Bis(imidazol-2-yl)me hane (2-BIM) and i s de i a i es (Fig. 1), in which he wo imidazole ings a e linked ia a single e ahed al ca bon a om, can be iewed as p imi i e models o mul i-his idine coo dina ion in biological sys ems. Such bis(imidazole) ligands ha e been shown o o m s able six-membe ed chela e ings wi h a a ie y o ansi ion me als [6–12], and he P (II) complex [P (2- BIM(Me)OH)Cl2] (2-BIM(Me)OH ¼bis(N-me hylim- idazol-2-yl)ca binol) [10] has been epo ed o exhibi no able an i umo ac i i y. 2. Resul s and discussion In he cu en s udy, AgClO4was eac ed a oom empe a u e wi h me hanolic solu ions o he fi e bis(imidazole) ligands shown in Fig. 1 o gi e he e- spec i e Ag(I) complex in mode a e o good yield. The complexes we e soluble in ho ace oni ile and DMSO and insoluble in wa e . The X- ay c ys al s uc u e o [Ag2(2-BIM)2](ClO4)2(Fig. 2 and Table 1) showed he complex o be cen osymme ic, con aining wo Ag(2- BIM)þuni s. The wo me al ions a e wo-coo dina e and ha e iden ical a oms in he plane o he chela ing ligand. Using i s wo imine N a oms each 2-BIM ligand b idges he pai o Ag(I) ions. All ou N a oms (N1, N4A, N4, N1A) a e coplana and he wo imidazole g oups in each 2-BIM ligand ha e an in e plana angle o 75.52(8)°. The wo Ag(I) ions a e only weakly in e - ac ing, he dis ance sepa a ing hem being 3.2612(4)  A. The binding o he wo 2-BIM ligands is simila o ha * Co esponding au ho . Tel.: +353-1-708-3767; ax: +353-1-708- 3815. E-mail add ess: [email p o ec ed] (M. McCann). 0277-5387/$ - see on ma e Ó2004 Else ie L d. All igh s ese ed. doi:10.1016/j.poly.2004.02.006 Polyhed on 23 (2004) 1249–1255 www.else ie .com/loca e/poly obse ed o he Biim2ligand (Biim2¼2,20-biimidaz- ola e) in he he e opolynuclea complex, [{Ru(pap)2 (Biim)}2Ag2](ClO4)2H2O (pap ¼2-(phenylazo)py i- dine) [13]. The sho e Ag–Ag sepa a ion in he la e species is (2.8899  A) is mainly a consequence o he smalle size o he Biim2ligand compa ed o 2-BIM (no me hylene b idge be ween he wo imidazole ings in Biim2). The o mula uni o [Ag2(2-BIM)2](ClO4)2is comple ed by wo la ice pe chlo a e, which a e in ol ed in hyd ogen bonding o he hyd ogen a oms o he im- idazole ni ogen a oms (N2 and N3, and N2A and N3A, espec i ely). The la ice pe chlo a e anions show un- symme ic b idging in e ac ions wi h Ag(I) ions o neighbou ing ca ions (AgOClO32.965(2) and 3.264(2)  A). The e a e hyd ogen bonding in e ac ions along he a and bdi ec ions which link he uni s oge he in he la ice, o ming shee s. Howe e , he e a e no hyd ogen bonds in he cdi ec ion, ha is o say, be ween he shee s. The e a e some in e molecula p–pin e ac ions in ol ing he imidazole ings on adjacen dime mole- cules and he e is also e idence o some in e dime C–Hpin e ac ions be ween neighbou ing imidazole unc ions. A pe spec i e iew o he cell packing is shown in Fig. 3. The X- ay c ys al s uc u e o [Ag2(2-BIM(Bz)- OH)2](ClO4)2E OH (Fig. 4 and Table 2) shows he asymme ic uni con aining one [Ag2(2-BIM(Bz)- OH)2]2þuni , wo pe chlo a e anions and a sol a e e hanol. The wo Ag(I) ions a e again b idged by he wo ligands and each me al is app oxima ely linea ly coo dina ed o a pai o imine ni ogens. The e a e ad- di ional longe in e ac ions wi h he pe chlo a e anions. The me al-me al dis ance o 3.018(4)  A is conside ably sho e han in [Ag2(2-BIM)2](ClO4)2. Wi hin he [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH complex molecule he e appea s o be some deg ee o in amolecula p–p in e ac ion be ween he imidazole and phenyl ings o he opposing wo 2-BIM(Bz)OH ligands in he dime . In e es ingly, he angles N(1A)–Ag(1)–Ag(2) and N(4B)–Ag(1)–Ag(2) a e e y diffe en o one ano he (168.30(12)°and 68.35(8)°, espec i ely). One o he pe chlo a e anions in he complex is diso de ed and was modeled wi h wo al e na i e se s o oxygen a om po- si ions (50% occupancy each). The e hanol sol a e is also diso de ed and was modeled as 50% occupancy o wo al e na i e posi ions. Hyd ogen a oms a ached o ca bon we e inse ed a calcula ed posi ions, whils hose a ached o he 2-BIM(Bz)OH alcohol g oups we e loca ed om diffe ence maps and no u he e- fined, hose bonded o he diso de ed oxygen a om o he e hanol sol a e we e no loca ed o included in he model. One iew o he packing in he c ys al is illus- a ed in Fig. 5. Repea ed a emp s o g ow c ys als o [Ag2{2-BIM (Me)}2](ClO4)2, [Ag2{2-BIM(CN)}2](ClO4)2and [Ag2{2- BIM(BzCl)}2](ClO4)2sui able o X- ay s uc u al anal- Fig. 2. X- ay c ys al s uc u e o [Ag2(2-BIM)2](ClO4)2. RR' 2-BIM H H 2-BIM(Bz)OH PhCH2 OH 2-BIM(Me) Me H 2-BIM(CN) NC(CH2)2H 2-BIM(ClBz) Cl2PhCH2 H N N N N HR' R R Fig. 1. Ligand symbols and s uc u es. Table 1 Selec ed bond leng hs (  A) and angles (°) o [Ag2(2-BIM)2](ClO4)2 Bond leng hs (  A) Ag(1)–N(4)#1 2.0953(17) Ag(1)–N(1) 2.0957(17) Ag(1)–Ag(1)#1 3.2612(4) Ag(1)–O(11)#2 2.965(2) Ag(1)–O(11)#3 3.264(2) Bond angles (°) N(4)#1–Ag(1)–N(1) 167.80(7) N(4)#1–Ag(1)–Ag(1)#1 91.08(5) N(1)–Ag(1)–Ag(1)#1 99.94(5) Symme y ans o ma ions used o gene a e equi alen a oms: #1 xþ1;yþ1;zþ1; #2 1 þx;1þy;z;#31x;y;1z. 1250 S. Abuskhuna e al. / Polyhed on 23 (2004) 1249–1255 ysis we e unsuccess ul. Howe e , gi en he close simila - i ies in he co e s uc u es o all fi e bis(imidazole) ligands he la e h ee sil e complexes a e hough o be essen- ially isos uc u al wi h [Ag2(2-BIM)2] (ClO4)2and [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH and a e hus also o mula ed as dime s. Fo [Ag2{2-BIM(CN)}2](ClO4)2 he e is also he possibili y o he pendan cyano g oups pa icipa ing in he bonding o he me al since Ag(I) has a high affini y o o ganoni ile dono s. 2.1. In i o an i-Candida s udies All o he me al- ee bis(imidazole) ligands and he Ag(I) complexes we e sc eened o hei abili y o inhibi he g ow h o C. albicans. The minimum inhibi o y concen a ion (MIC) is he concen a ion o d ug (ex- p essed as lg o complex pe 1 cm3o aqueous g ow h medium solu ion) equi ed o o ally inhibi he g ow h o he ungal cells a 37 °C. As he complexes and he Fig. 4. X- ay c ys al s uc u e o [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH. Fig. 3. Packing diag am o [Ag2(2-BIM)2](ClO4)2. S. Abuskhuna e al. / Polyhed on 23 (2004) 1249–1255 1251 ligands we e essen ially wa e insoluble he es s we e ca ied ou as suspensions o he complexes in he aqueous g ow h medium. In addi ion, because he complexes we e soluble in DMSO a se ies o an i- ungal es s we e also conduc ed using DMSO/wa e solu ions o he complexes (see Sec ion 3). In all cases, he ac i i y o he complexes we e compa ed o ha o a con ol sys em in which he cells we e g own in he absence o any added complex o ee ligand (unde hese condi- ions he ungal cell eplica ion was apid). In addi ion, unde he p esen biological es condi ions an aqueous suspension o he an i-Candida p esc ip ion d ug ke o- conazole had an MIC alue o ca. 2.5 lgcm 3. All o he me al- ee ligands we e ound o be inac i e agains he ungal cells. As aqueous suspensions, he sil e complexes we e mode a ely ac i e wi h MIC alue anges indica ed in b acke s: [Ag2(2-BIM)2](ClO4)2(10– 20 lgcm 3); [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH (50– 100 lgcm 3); [Ag2{2-BIM(Me)}2](ClO4)2(20–50 lgcm 3); [Ag2{2-BIM(CN)}2](ClO4)2(20–50 lgcm 3). A conside able imp o emen in an i- ungal ac i i y was obse ed upon adminis e ing he complexes as DMSO/ wa e solu ions as opposed o aqueous suspensions: [Ag2(2-BIM)2](ClO4)2(5–10 lgcm 3); [Ag2(2-BIM(Bz)- OH)2](ClO4)2E OH (2.5–5 lgcm 3); [Ag2{2-BIM (Me)}2](ClO4)2(2.5–5 lgcm 3); [Ag2{2-BIM(CN)}2] (ClO4)2(5–10 lgcm 3); [Ag2{2-BIM(BzCl)}2](ClO4)2 (5–10 lgcm 3). Besides g ea ly inc easing he solubili y o he complexes he DMSO is also hough o make he cells mo e pe meable o he complexes so ha hey can en e he cells and exe hei effec s on he o ganism. In compa ison, as an aqueous suspension [Ag(phen- dio)2]ClO4has an MIC alue o 0.3 lgcm 3[5]. 3. Expe imen al Chemicals we e pu chased om comme cial sou ces and, unless specified, we e used wi hou u he pu ifi- ca ion. Li e a u e me hods we e used o p epa e 2-BIM [14,15], 2-BIM(Me) [7,16] and 2-BIM(OH)Bz [17]. The p epa a ion o he sil e complexes was conduc ed in he absence o ligh and he samples we e s o ed in he da k. In a ed spec a o solids (in a KB ma ix) we e e- co ded in he egion 4000–400 cm1on a Nicole FT-IR Impac 400D in a ed spec ome e and 1H NMR spec a we e un on a B uke A ance 300 MHz ins u- men . Mic oanaly ical da a we e p o ided by he Mi- c oanaly ical Labo a o y, Na ional Uni e si y o I eland, Co k, I eland. 3.1. 2-BIM(CN) Ac yloni ile (3.6 g, 67 mmol) was added o a mix u e o 2-BIM (5.0 g, 34 mmol), and ie hylamine (6.8 g, 67 mmol) in ace oni ile (150 cm3). The eac ion mix u e was efluxed wi h s i ing o 5 h. The sol en was e- mo ed unde educed p essu e gi ing a whi e solid, which was ec ys allised om ace one. Yield: 4.7 g (55%). Mp 170 °C. Anal. Calc. C, 61.40; H, 5.50; N, Table 2 Selec ed bond leng hs (  A) and angles (°) o [Ag2(2-BIM(Bz)OH)2] (ClO4)2E OH Bond leng hs (  A) Ag(1)–N(1A) 2.111(3) Ag(1)–N(4B) 2.134(3) Ag(1)–Ag(2) 3.0184(4) Ag(1)–O(11) 3.211(6) Ag(1)–O(13) 3.312(7) Ag(2)–N(1B) 2.119(3) Ag(2)–N(4A) 2.141(3) Ag(2)–O(21) 3.012(7) Ag(2)–O(210) 3.234(14) Bond angles (°) N(1A)–Ag(1)–N(4B) 168.30(12) N(1A)–Ag(1)–Ag(2) 103.65(8) N(4B)–Ag(1)–Ag(2) 68.35(8) N(1A)–Ag(1)–O(11) 82.69(13) N(4B)–Ag(1)–O(11) 105.55(13) Ag(2)–Ag(1)–O(11) 173.58(10) N(1A)–Ag(1)–O(13) 109.69(14) N(4B)–Ag(1)–O(13) 81.74(14) Ag(2)–Ag(1)–O(13) 133.53(10) O(11)–Ag(1)–O(13) 41.35(13) N(1B)–Ag(2)–N(4A) 172.79(12) N(1B)–Ag(2)–O(21) 92.53(19) N(4A)–Ag(2)–O(21) 93.9(2) N(1B)–Ag(2)–Ag(1) 99.97(8) N(4A)–Ag(2)–Ag(1) 74.23(8) O(21)–Ag(2)–Ag(1) 164.88(17) N(1B)–Ag(2)–O(210) 76.8(2) N(4A)–Ag(2)–O(210) 109.3(2) Ag(1)–Ag(2)–O(210) 174.4(2) Fig. 5. Packing diag am o [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH. 1252 S. Abuskhuna e al. / Polyhed on 23 (2004) 1249–1255 33.07. Found: C, 61.26; H, 5.47; N, 32.76%. 1H NMR(DMSO-d6): 7.17 (2H, d, J¼1:3 Hz), 6.8 (2H, d, J¼1:3 Hz), 4.3 (4H, , J¼6:7 Hz), 4.2 (2H, s), 3.0 (4H, , J¼6:7 Hz) ppm. 13C NMR(DMSO-d6): 143.6, 127.3, 120.8, 118.8, 25.4, 19.2 ppm. IR: 3125, 2932, 2264, 1485, 1447, 1383, 1287, 1166, 1134, 1089, 926, 722, 743, 705 cm1. Mass spec.: m=z: 254 (Mþ, 4%), 200 (4), 173 (8), 161 (6), 148 (10), 134 (11), 120 (14), 107 (14), 94 (28), 82 (84), 66 (20), 54 (100). 3.2. 2-BIM(BzCl) 2,6-Dichlo obenzyl chlo ide (0.53 g, 2.7 mmol) was added o a s i ed mix u e o 2-BIM (0.2 g, 1.3 mmol) and sodium hyd ide (0.06 g, 2.5 mmol) in d y DMF (15 cm3). The mix u e was s i ed o e nigh a oom em- pe a u e and he sol en emo ed unde educed p es- su e. Wa e (20 cm3) was added and he p oduc ex ac ed wi h dichlo ome hane, and he dichlo ome- hane solu ion was hen d ied o e magnesium sulpha e. A e fil a ion he sol en was emo ed unde educed p essu e. The esul ing b own oily esidue was diges ed in cyclohexane and he whi e p oduc p ecipi a ed upon cooling. Yield: 0.34 g (54%). Mp 93 °C. Anal. Calc. C, 54.10; H, 3.43; N, 12.02. Found: C, 54.09; H, 3.58; N, 11.73%. 1H NMR (CDCl3): 7.4 (2H, d, J¼2 Hz), 7.1 (2H, dd, J¼8:3 Hz), 6.9 (2H, d, J¼1:3 Hz), 6.7 (2H, d, J¼1:3 Hz), 6.5 (2H, d, J¼8:3 Hz), 5.2 (4H, s), 4.2 (2H, s). 13C NMR (CDCl3): 143, 134.5, 133.6, 132.7, 129.5, 129.1, 128.1, 127.5, 120.5, 47, 27 ppm. IR: 2945, 1593, 1485, 1447, 1389, 1274, 1121, 1057, 849, 733, 682 cm1. Mass spec.: m=z: 466 (Mþ, 16%), 429 (4), 305 (26), 269 (8), 205 (8), 159 (100), 147 (8), 123 (16), 99 (6), 89 (16), 81 (38), 63 (10). 3.3. [Ag2(2-BIM)2](ClO4)2 Sil e pe chlo a e (0.70 g, 3.37 mmol) was added o a s i ed solu ion o 2-BIM (0.5 g, 3.37 mmol) in me hanol (15 cm3) and he mix u e s i ed o 1 h a oom em- pe a u e. The p ecipi a ed whi e was fil e ed off, washed wi h me hanol and ai -d ied. Yield: 1.00 g (83%). Anal. Calc. C, 23.64; H, 2.25; N, 15.76. Found: C, 23.58; H, 2.13; N, 15.78%. IR: 3344, 3151, 1293, 1095, 759, 624 cm1.1H NMR (ppm DMSO): 6.8 (8H, s), 4 (4H, s). The complex was soluble in ho ace oni ile and DMSO, and c ys als sui able o X- ay analysis we e ob ained by ec ys allising om ace oni ile. 3.4. [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH This whi e complex was p epa ed and ec ys allised in a simila way o [Ag2(2-BIM)2](ClO4)2using sil e pe - chlo a e (0.06 g, 0.17 mmol) and 2-BIM(Bz)OH (0.1 g, 0.29 mmol). Yield: 0.11 g (68%). Anal. Calc. C, 43.87; H, 3.48; N, 9.70. Found: C, 44.21; H, 3.45; N, 9.69%. IR: 2485, 3138, 2971, 1497, 1459, 1287, 1102, 827, 717, 624 cm1.1H NMR (ppm DMSO): 5.2 (8H, dd), 6.4 (2H, d), 7.1 (8H, s), 7.35 (20H, m). The complex was soluble in ho ace oni ile, ho ace one and DMSO. 3.5. [Ag2 { 2-BIM(Me) } 2](ClO4)2 To a solu ion o sil e pe chlo a e (0.10 g, 0.48 mmol) in me hanol (3 cm3) was added a solu ion o 2-BIM(Me) (0.10 g, 0.56 mmol) in me hanol (3 cm3). A whi e p e- cipi a e o med immedia ely and he eac ion mix u e was s i ed o 20 min a oom empe a u e. The whi e p oduc was fil e ed off, washed wi h me hanol and hen d ied in ai . Yield: 0.13 g (62%). Anal. Calc. C, 28.10; H, 3.13; N, 14.60. Found: C, 27.5; H, 3.00; N, 14.13%. IR: 3485, 3138, 2971, 1497, 1414, 1293, 1102, 772, 627 cm1. 1H NMR (ppm DMSO): 3.5 (12H, s), 4.36 (4H, s), 6.8 (4H, s), 7.2 (4H, s). The complex was soluble in ho ace oni ile, ho ace one and DMSO. 3.6. [Ag2 { 2-BIM(CN) } 2](ClO4)2 This whi e complex was p epa ed in a simila way o [Ag2{2-BIM(Me)}2](ClO4)2using sil e pe chlo a e (0.10 g, 0.48 mmol) and 2-BIM(CN) (0.20 g, 0.78 mmol). Yield: 0.32 g (89%). Anal. Calc. C, 33.81; H, 3.03; N, 18.20. Found: C, 33.46; H, 2.92; N, 18.20%. IR: 3511, 3138, 2984, 2264, 1497, 1427, 1287, 1102, 756, 627 cm1. 1H NMR (ppm DMSO): 7.3 (4H, s), 6.8 (4H, s), 4.4 (4H, s), 4.2 (8H, ), 2.9 (8H, ). The complex was soluble in ho ace oni ile, ho ace one and DMSO. 3.7. [Ag2 { 2-BIM(BzCl) } 2](ClO4)2 This whi e complex was p epa ed in a simila way o [Ag2{2-BIM(Me)}2](ClO4)2using sil e pe chlo a e (0.08 g, 0.38 mmol) in 3 ml me hanol was added o 2- BIM(BzCl) (0.20 g, 0.43 mmol). Yield: 0.16 g (57%). Anal. Calc. C, 37.43; H, 2.37; N, 8.31. Found: C, 37.03; H, 2.41; N, 8.04%. IR: 3511, 3138, 1593, 1491, 1395, 1274, 1102, 836, 749, 628 cm1.1H NMR (ppm DMSO): 7.9 (4H, d), 7.56 (4H, dd), 7.5 (4H, d),7.25 (4H, d) 6.8 (4H, d), 5.5 (8H, s), 4.8 (4H, s). The complex was soluble in ho ace oni ile, ho ace one and DMSO. 3.8. An i-Candida es ing Candida albicans ATCC 10231 was ob ained om he Ame ican Type Cul u e Collec ion (Manasas, VA, USA). Cul u es we e g own on Sabou aud dex ose aga (SDA) pla es a 37 °C and main ained a 4 °C o sho - e m s o age. Minimum inhibi o y concen a ions (MICs) we e de e mined as ollows. RPMI-1640 b o h medium was used o he an i-Candida suscep ibili y es ing. RPMI (5.15 g) was dissol ed in cold dis illed wa e (425 cm3) in a 1-l Du an bo le and he pH S. Abuskhuna e al. / Polyhed on 23 (2004) 1249–1255 1253 adjus ed o 4.0 using a ew d ops o HCl (ca. 2 M). The esul ing solu ion was au ocla ed and hen allowed o cool o app oxima ely 50 °C. Mo pholinep opanesul - onic acid (MOPS) (17.3 g) and LL -glu ama e (0.15 g) we e dissol ed oge he wi h s i ing in dis illed wa e (50 cm3) and he esul ing solu ion fil e s e ilized. The MOPS and LL -glu ama e solu ion was added o he wa m RPMI and he pH o he mix u e adjus ed o 7.0 using s e ile NaOH (6 M). P io o MIC es ing, cells we e g own on SDA a 37 °C o 24 h. Cell suspensions we e p epa ed in s e ile phospha e buffe ed saline (PBS, pH 7.2) and cells we e coun ed mic oscopically ollowing dilu ion wi h PBS. A mic o i e pla e was inocula ed wi h cells a a densi y o 5 105cells cm3. Tes sus- pensions (in wa e ) and solu ions (in aqueous DMSO) o he sil e complexes we e each p epa ed as ollows. Suspensions in wa e : he complex (0.02 g) was sus- pended in dis illed wa e (10 cm3) o yield a s ock sus- pension o concen a ion 2000 lgcm 3. Doubling dilu ions o he s ock suspension we e made o yield a se ies o es suspensions anging in complex concen- a ion om 20 o 1.25 lgcm 3.Solu ions in DMSO: he complex (0.02 g) was dissol ed in DMSO (10 cm3) o yield a s ock solu ion o concen a ion 2000 lgcm 3. Doubling dilu ions o he s ock solu ion we e made using dis illed wa e o yield a se ies o es solu ions anging in complex concen a ion om 20 o 1.25 lgcm 3. The DMSO concen a ion (% / ) in he es solu ions anged om 1.0% o 0.06% (no e ha a 2% DMSO aqueous solu ion, wi hou added complex, does no inhibi he g ow h o he mic oo ganism whe eas a 3% DMSO aqueous solu ion will a es cell g ow h o- ally). Fo he aqueous es suspensions and he aque- ous-DMSO es solu ions he complex/cell mix u es we e incuba ed a 37 °C o 24 h wi h con inuous shaking and he assays we e pe o med in iplica e. Pla es we e ead using a Labsys ems iEMS Reade MF (abso bance a k¼540 nm) and da a we e s a is ically analysed. 3.9. X- ay c ys allog aphy All o he da a we e collec ed a 150 K on a B uke SMART 1000 diff ac ome e using Mo Ka adia ion (k¼0:71073  A). The s uc u es we e sol ed by di ec me hods and efined by ull-ma ix leas -squa es on F2 using all he eflec ions. One o he p echlo a e anions in [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH is diso de ed and was modeled wi h wo al e na i e se s o oxygen a om posi ions (50% occupancy each). The e hanol sol a e is also diso de ed and was modeled as 50% occupancy o wo al e na i e posi ions. All non-hyd ogen a oms we e efined wi h aniso opic a omic displacemen pa ame- e s and hyd ogen a oms bonded o ca bon we e in- se ed a calcula ed posi ions. Hyd ogen a oms bonded o ni ogen ([Ag2(2-BIM)2](ClO4)2) o he ligand alco- hol g oup ([Ag2(2-BIM(Bz)OH)2](ClO4)2E OH) we e loca ed om diffe ence maps and no u he efined; hose bonded o he diso de ed oxygen o he e hanol sol a e in [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH we e no included in he model. All p og ammes used in he s uc u e solu ion and efinemen a e included in he SHELXTLSHELXTL package [18] and da a collec ion and efine- men de ails a e summa ised in Table 3. Table 3 X- ay c ys allog aphic da a. C ys al da a and s uc u e efinemen o [Ag2(2-BIM)2](ClO4)2and [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH Iden ifica ion code [Ag2(2-BIM)2](ClO4)2[Ag2(2-BIM(Bz)OH)2](ClO4)2E OH Empi ical o mula C14H16Ag2Cl2N8O8C44H46Ag2Cl2N8O11 Fo mula weigh 710.99 1149.53 C ys al sys em iclinic iclinic Space g oup P  1P  1 a(  A) 7.8149(10) 10.9636(7) b(  A) 8.9902(11) 14.7743(9) c(  A) 9.2559(11) 16.0180(10) a(°) 108.046(2) 114.3870(10) b(°) 98.283(2) 98.2900(10) c(°) 114.763(2) 95.8410(10) Volume (  A3) 532.47(11) 2300.6(2) Z12 F(0 0 0) 348 1164 C ys al size (mm) 0.16 0.25 0.31 0.37 0.25 0.14 h ange o da a collec ion (°) 2.44–28.65 1.54–25.00 Reflec ions collec ed 4544 16522 Independen eflec ions [Rðin Þ] 2396 [0.0186] 8050 [0.0159] Abso p ion co ec ion mul iscan mul iscan Maximum and minimum ansmission 1.0000 and 0.8116 1.000000 and 0.912980 Da a/ es ain s/pa ame e s 2396/0/154 8050/98/667 R1,wR2[I>2 ðIÞ] 0.0198, 0.0526 0.0356, 0.0877 R1,wR2(all da a) 0.0204, 0.0530 0.0455, 0.0942 1254 S. Abuskhuna e al. / Polyhed on 23 (2004) 1249–1255 4. Supplemen a y da a X- ay supplemen a y da a o [Ag2(2-BIM)2](ClO4)2 and [Ag2(2-BIM(Bz)OH)2](ClO4)2E OH a e a ailable om he Camb idge C ys allog aphic Da a Cen e, 12 Union Road, Camb idge CB2 1EZ, England ( ax: +44- 1223-336033; E-mail: deposi @ccdc.cam.ac.uk o www: h p://www.ccdc.cam.ac.uk), on eques quo ing he de- posi ion numbe s CCDC 219908 and CCDC 219909, espec i ely. Acknowledgemen s S. Abuskhuna acknowledges financial suppo om he Libyan Highe Educa ion Depa men . Re e ences [1] B. Coyle, K. Ka anagh, M. McCann, M. De e eux, M. Ge agh y, BioMe als 16 (2003) 321. [2] B. Coyle, P. Kinsella, M. McCann, M. De e eux, R. O’Conno , M. Clynes, K. Ka anagh, Toxicology in Vi o 18 (2004) 63. [3] M. Ge agh y, J.F. C onin, M. De e eux, M. McCann, BioMe als 13 (2000) 1. [4] M. McCann, B. Coyle, J. B iody, F. Bass, N. O’Go man, M. De e eux, K. Ka anagh, V. McKee, Polyhed on 22 (2003) 1595. [5] M. McCann, B. Coyle, S. McKay, P. McCo mack, K. 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