Complexes against Pathogenic Bacteria, Including Methicillin

Sep 22, 2016 - While methicillin-resistant Staphylococcus aureus (MRSA) is the most well-known, a wide range of other drug resistant pathogenic bacter...
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Antimicrobial Properties of Tris(homoleptic) Ruthenium(II) 2‑Pyridyl1,2,3-triazole “Click” Complexes against Pathogenic Bacteria, Including Methicillin-Resistant Staphylococcus aureus (MRSA) Sreedhar V. Kumar,†,‡,§ Synøve Ø. Scottwell,†,‡,§ Emily Waugh,‡ C. John McAdam,† Lyall R. Hanton,† Heather J. L. Brooks,‡ and James D. Crowley*,† †

Department of Chemistry, University of Otago, PO Box 56, Dunedin, New Zealand Department of Microbiology and Immunology, Otago School of Medical Sciences, University of Otago, PO Box 56, Dunedin, New Zealand

‡

S Supporting Information *

ABSTRACT: A series of tris(homoleptic) ruthenium(II) complexes of 2-(1-R-1H-1,2,3-triazol-4-yl)pyridine “click” ligands (R-pytri) with various aliphatic (R = butyl, hexyl, octyl, dodecyl, and hexdecyl) and aromatic (R = phenyl and benzyl) substituents was synthesized in good yields (52%− 66%). The [Ru(R-pytri)3]2+(X−)2 complexes (where X− = PF6− or Cl−) were characterized by elemental analysis, highresolution electrospray ionization mass spectrometry (HR-ESIMS), 1H and 13C nuclear magnetic resonance (NMR) and infrared (IR) spectroscopies, and the molecular structures of six of the compounds confirmed by X-ray crystallography. 1H NMR analysis showed that the as-synthesized materials were a statistical mixture of the mer- and fac-[Ru(R-pytri)3]2+ complexes. These diastereomers were separated using column chromatography. The electronic structures of the mer- and fac-[Ru(Rpytri)3]2+ complexes were examined using ultraviolet−visible (UV-Vis) spectroscopy and cyclic and differential pulse voltammetry. The family of R-pytri ligands and the corresponding mer- and fac-[Ru(R-pytri)3]2+ complexes were tested for antimicrobial activity in vitro against both Staphylococcus aureus and Escherichia coli bacteria. Agar-based disk diffusion assays indicated that two of the [Ru(R-pytri)3](X)2 complexes (where X = PF6− and R = hexyl or octyl) displayed good antimicrobial activity against Gram-positive S. aureus and no activity against Gram-negative E. coli at the concentrations tested. The most active [Ru(R-pytri)3]2+ complexes ([Ru(hexpytri)3]2+ and Ru(octpytri)3]2+) were converted to the water-soluble chloride salts and screened for their activity against a wider range of pathogenic bacteria. As with the preliminary screen, the complexes showed good activity against a variety of Gram-positive strains (minimum inhibitory concentration (MIC) = 1−8 μg/mL) but were less effective against Gram-negative bacteria (MIC = 16−128 μg/mL). Most interestingly, in some cases, the ruthenium(II) “click” complexes proved more active (MIC = 4−8 μg/mL) than the gentamicin control (MIC = 16 μg/mL) against two strains of methicillin-resistant S. aureus (MRSA) (MR 4393 and MR 4549). Transmission electron microscopy (TEM) experiments and propidium iodide assays suggested that the main mode of action for the ruthenium(II) R-pytri complexes was cell wall/ cytoplasmic membrane disruption. Cytotoxicity experiments on human dermal keratinocyte and Vero (African green monkey kidney epithelial) cell lines suggested that the complexes were only modestly cytotoxic at concentrations well above the MIC values.

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MRSA, fluoroquinolone-resistant P. aeruginosa, and vancomycin-resistant Enterococcus (VRE) are a serious cause of concern in hospitals, because of rapidly increasing rates of infection.2 Clearly there is a pressing need for new effective antimicrobial agents with novel modes of action. Metal complexes3 are promising in this regard, when compared to traditional organic antimicrobial drugs, as they offer alternative electronic and stereochemical properties, which may lead to new modes of action.4 Over 60 years ago, Dwyer

INTRODUCTION The continued emergence of drug-resistant bacteria, coupled with a lack of new antimicrobial drugs in the pharmaceutical pipeline, is one of the most serious health issues of the 21st century. A recent World Health Organization (WHO) report1 has suggested that the world is on the cusp of a post-antibiotic era. While methicillin-resistant Staphylococcus aureus (MRSA) is the most well-known, a wide range of other drug resistant pathogenic bacteria have now been identified including Mycobacterium tuberculosis, Pseudomonas aeruginosa, Enterococcus faecium, Klebsiella pneumoniae, Acinetobacter baumanii, Neisseria gonorrheae, and Enterobacter species.1 In particular, © XXXX American Chemical Society

Received: July 1, 2016

A

DOI: 10.1021/acs.inorgchem.6b01574 Inorg. Chem. XXXX, XXX, XXX−XXX

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Inorganic Chemistry and co-workers5 showed that inert lipophilic ruthenium(II) complexes of the N−N bidentate chelators 2,2-bipyridine (bipy) and 1,10-phenanthroline (phen) (e.g., 1) were effective antimicrobial agents (Figure 1).4c The need for new classes of

Scheme 1. Synthesis of the Ruthenium(II) R-pytri Complexes, [Ru(R-pytri)3](PF6)2a

a Conditions: (i) (a) R-pytri (3 equiv), RuCl3·nH2O (1 equiv), ethanol−water (7:3), microwave irradiation (220 W), 120 °C, 40 min or 1 h, and (b) aqueous NH4PF6 (5 equiv).

photophysical,12 and catalytic13 properties of these complexes have been examined and di-,14 tri-,15 and tetra-topic16 pytri ligands have been exploited to synthesize metallosupramolecular systems.14a,16 Due to the facile functionalization of these “click” ligands, there has been considerable interest in the use of their complexes in a biological context. Functionalized R-pytri complexes of inert octahedral ions, such as Re(I),17 Tc(I),17b,i,j Ru(II),18 and Ir(III),18,19 have been exploited as infrared, luminescent, and radiolabeled bioprobes. In addition, square planar (Pt(II) and Pd(II)) and octahedral (Re(I), and Ir(III)) R-pytri complexes have displayed good cytotoxicity against some cancer cell lines.17d,g,h,j,19b,20 We have previously examined the use of monometallic and dimetallic R-pytri complexes as antibacterial21 and antifungal22 agents with modest success. Herein, we report the synthesis of a series of tris(homoleptic) ruthenium(II) complexes of the R-pytri “click” ligands. The antibacterial properties of ligands and complexes were screened against S. aureus and E. coli. The most active complexes were converted to the water-soluble chloride salts and screened for their activity against a wider range of pathogenic bacteria. In addition, the biological mode of action of the complexes was examined using transmission electron microscopy (TEM) experiments and propidium iodide assays.

Figure 1. Schematics of inert ruthenium(II) polypyridyl complexes, which display antimicrobial activity.

antimicrobial agents has reinvigorated research into these systems. Keene and co-workers investigated the antimicrobial activity of both mononuclear and dinuclear ruthenium(II) complexes against S. aureus and MRSA.6 The parent [Ru(phen)3]2+ complex, 1, displayed very modest activity against both S. aureus and MRSA (minimum inhibitory concentration (MIC) of >128 μg/mL) but the more lipophilic [Ru(Me4phen)3]2+ complex, 2, was highly active (S. aureus, MIC = 0.5 μg/mL and MRSA, MIC = 4 μg/mL).6 The related ruthenium(II) complexes 3 (MIC = 2−8 μg/mL)7 and 4 (MIC = 6.25 μg/mL)8 were also effective against various strains of MRSA (Figure 1). However, to date, the most active ruthenium(II)-based antimicrobial agents are the dinuclear complexes, 5 (n = 8, 10, or 12), developed by Keene and coworkers (Figure 1).6 These systems have high activity against both Gram-positive (S. aureus and MRSA, MIC = 1 μg/mL) and Gram-negative bacteria (P. aeruginosa and Escherichia coli (E. coli), MIC = 1 μg/mL). Recently 2-(1-R-1H-1,2,3-triazol-4-yl)pyridine “click” ligands (R-pytri, Scheme 1) have emerged as readily functionalized analogues of the ubiquitous 2,2′-bipyridine/1,10-phenanthroline N−N chelators.9 Because these ligands are generated using the functional-group-tolerant copper(I)-catalyzed azide alkyne cycloaddition (CuAAC, or “click”) reaction,10 an extremely diverse array of functionalized ligands and their corresponding complexes have been generated.9 The electrochemical,11

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RESULTS AND DISCUSSION Ligand Synthesis. The 2-(1-R-1H-1,2,3-triazol-4-yl)pyridine “click” ligands (Bnpytri, Phpytri, and octpytri; see Scheme 1) were synthesized using our previously reported procedures.23 Similarly, the new ligands (butpytri, hexpytri, dodecpytri, and hexdecpytri; see Scheme 1) were generated by mixing 2-ethynylpyridine, the appropriate alkyl bromide, NaN3, CuSO4·5H2O, and sodium ascorbate in DMF/H2O (4:1) and heating at 95 °C for 20 h.15,23 This method provided, after workup, the desired compounds in good to excellent yields (65%−90%) without the need for isolating the potentially hazardous azide intermediates. Each of the ligands were characterized using 1H, 13C NMR, and IR spectroscopies, high-resolution electrospray ionization mass spectrometry (HRESI-MS), and elemental analysis (Section 2 (Figures S1−S8) in the Supporting Information) and the collected data were completely consistent with the expected structures. B

DOI: 10.1021/acs.inorgchem.6b01574 Inorg. Chem. XXXX, XXX, XXX−XXX

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Inorganic Chemistry Synthesis of the Ruthenium(II) R-pytri Complexes, [Ru(R-pytri)3]2+. The groups of Fletcher24 and Schubert25 have previously reported the synthesis of some [Ru(R-pytri)3]2+ (X−)2 complexes, where X− was either Cl− or PF6−. While both these methods lead to the formation of the [Ru(R-pytri)3]2+ complexes, the microwave-enhanced conditions previously used by Gunnlaugsson and co-workers to generate some related [Ru(R-tripy)2]2+ (where R-tripy = 2,6-bis(1,2,3-triazol-4-yl)pyridine) provided the most facile access to the desired family of compounds.26 One of the R-pytri ligands (3 equiv) and RuCl3·nH2O (1 equiv) were dissolved in an ethanol−water mixture (7:3) and irradiated with microwaves (220 W) at 120 °C for either 40 min or 1 h. Under these reaction conditions, the Ru(III) was reduced to Ru(II) and the addition of an aqueous solution of ammonium hexafluorophosphate (NH4PF6, 5 equiv) resulted in the precipitation of the yellow/orange [Ru(R-pytri)3](PF6)2 complexes in good yields (52%−66%). The presence of both ligand (3000−2900 cm−1, C−H stretch) and the PF6− counteranion (839−828 cm−1, P− F stretch) in the isolated yellow/orange solids was confirmed by IR spectroscopy, and elemental analyses were consistent with the expected [Ru(R-pytri)3](PF6)2 formulations. Highresolution electrospray ionization mass spectra (acetonitrile or acetone) of the isolated yellow/orange materials displayed isotopically resolved peaks consistent with the presence of [Ru(R-pytri)3]2+ and [Ru(R-pytri)3(PF6)]+ ions for each complex, confirming the formation of the desired complexes (Section 3 (Figures S9 and S10) in the Supporting Information). 1H NMR analysis (CD3CN) showed, as expected,24,25 that the isolated materials were a statistical mixture of the mer- and fac-[Ru(R-pytri)3]2+ (Figure 2b). A diagnostic24,25 downfield shift of the triazole proton He is observed upon complexation to the ruthenium(II) ion (Figure 2b). The as-synthesized material displays four triazole resonances (three corresponding to the mer-[Ru(R-pytri)3]2+ diastereomer and the additional peak from the highersymmetry fac-[Ru(R-pytri)3]2+ diastereomer.24,25 These diastereomers were painstakingly separated using column chromatography (silica gel) to give the pure mer-[Ru(Rpytri)3]2+ and fac-[Ru(R-pytri)3]2+ (see Figures 2c and 2d, as well as Section 3 (Figures S11−S44) in the Supporting Information). Although the mer and fac diastereomers were separated, we have not attempted to resolve the Λ and Δ enantiomers of the chiral cations, thus the mer- and fac-[Ru(Rpytri)3]2+ complexes are racemic (rac) mixtures. Vapor diffusion between either acetonitrile or acetone solutions of the hexafluorophosphate complexes (fac-[Ru(Phpytri)3]2+, fac-[Ru(Bnpytri)3]2+, fac-[Ru(hexpytri)3]2+, mer-[Ru(Phpytri)3]2+, mer-[Ru(Bnpytri)3]2+, and mer-[Ru(hexpytri)3]2+) and diisopropyl ether produced crystals of sufficient quality for analysis by X-ray crystallography (see Figure 3, as well as Section 5 in the Supporting Information). As expected, the ruthenium(II) ions of the complexes are coordinated in an octahedral fashion to three bidentate R-pytri ligands with the divalent charge of the metal ion balanced by two noncoordinating PF6− anions (see Figure 3, as well as Section 5 in the Supporting Information). These tris-bidentate coordinated fac- and mer-[Ru(R-pytri)3]2+ cations are chiral and a racemic mixture of the Λ and Δ enantiomers is present in the crystals. The ruthenium−nitrogen bond distances (Ru− Npyridyl = 2.067−2.096 Å and Ru−Ntriazolyl = 2.014−2.068 Å) and bond angles (Figure 3) are similar to those previously

Figure 2. Partial 1H NMR (CD3CN, 298 K, 400 MHz) stacked plot of the (a) hexpytri ligand and (b−d) ruthenium(II) hexpytri complexes. Panel (b) indicates that the “as-synthesized” material is a mixture of mer- and fac-[Ru(hexpytri)3]2+ complexes, which can be separated into the pure fac-diastereomers (panel (c)) and mer-diastereomers (panel (d)), using column chromatography (silica gel).

observed11a,12d,13a,24,25,27 in related monomeric ruthenium(II) R-pytri complexes. Electronic Spectroscopy. The electronic absorption properties (CH3CN, 10−5 M) of the pure mer and fac diastereomers were essentially identical to those observed previously for mer/fac mixtures (see Figure 4, as well as Section 6 (Figures S45 and S46) in the Supporting Information).24,25 The [Ru(R-pytri)3](PF6)2 complexes (with alkyl and benzyl substituents) display two strong absorption bands, which have been attributed to ligand-centered (LC, 270−280 nm) and metal-to-ligand charge transfer (MLCT, 380−390 nm) transitions.24,25 The [Ru(Phpytri)3](PF6)2 complexes (mer and fac) showed an additional broad band between 300 and 340 nm, presumably due to the extended conjugation present in these compounds. The position of the MLCT (380−390 nm was essentially unaffected by the nature of the substituent on the R-pytri ligands (Figure 4). In addition, there was no obvious difference between the spectra of the pure mer and fac diastereomers for any of the complexes studied herein (see Figure S46). This behavior differs from that observed by Schubert and co-workers,25 who found that the MLCT bands for the mer and fac isomers of [Ru(Naphthalene-pytri)3](PF6)2 were different by 14 nm. Presumably, the larger size and extended conjugation in the Schubert compound leads to the observed differences. Compared to the related [Ru(bipy)3]2+ and [Ru(octpytri)(bipy)2]2+ complexes (vide inf ra, brown trace in Figure 4), the MLCT bands of the [Ru(R-pytri)3]2+ complexes are blue-shifted. This behavior has been observed C

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Figure 3. Perspective views showing the molecular structures of the ruthenium(II) R-pytri cations: (a) fac-[Ru(Bnpytri)3]2+, (b) mer[Ru(Bnpytri)3]2+, (c) fac-[Ru(Phpytri)3]2+, (d) mer-[Ru(Phpytri)3]2+, (e) fac-[Ru(hexpytri)3]2+, and (f) mer-[Ru(hexpytri)3]2+. Only the Δ enantiomer is shown for each complex. Hydrogen atoms, solvent molecules, and hexafluorophosphate counterions are omitted for the sake of clarity. Thermal ellipsoids are drawn at the 50% probability level. Selected bond lengths (Å) and angles (deg) are given in the text. For fac-[Ru(Bnpytri)3]2+ (Figure 3a): Ru1−N1, 2.089(1) Å; Ru1−N2, 2.026(2) Å; Ru1−N5, 2.089(2) Å; Ru1−N6, 2.029(1) Å; Ru1−N9, 2.096(2) Å; Ru1−N10, 2.019(2) Å; N1−Ru1−N2, 78.51(7) Å; N5−Ru1−N6, 78.31(7)°; N9−Ru1−N10, 77.97(7)°; N1−Ru1−N6, 172.55(7)°; N5−Ru1−N10, 175.99(7)°; and N9−Ru1−N2, 168.44(7)°. For mer-[Ru(Bnpytri)3]2+ (Figure 3b): Ru1−N1, 2.083(4) Å; Ru1−N2, 2.068(4) Å; Ru1−N5, 2.094(4) Å; Ru1−N6, 2.054(4) Å; Ru1−N9, 2.088(5) Å; Ru1−N10, 2.064(4) Å; N1−Ru1−N2, 78.4(2)°; N5−Ru1−N6, 78.6(2)°; N9−Ru1−N10, 78.7(2)°; N1−Ru1− N10, 172.0(2)°; N2−Ru1−N6, 168.0(2)°; N5−Ru1−N9, 175.2(2)°. For fac-[Ru(Phpytri)3]2+ (Figure 3c): Ru1−N1, 2.096(2) Å; Ru1−N2, 2.024(2) Å; Ru1−N5, 2.088(2) Å; Ru1−N6, 2.022(2) Å; Ru1−N9, 2.080(2) Å; Ru1−N10, 2.024(2) Å; N1−Ru1−N2, 78.17(8)°; N5−Ru1−N6, 78.28(8)°; N9−Ru1−N10, 78.57(8)°; N1−Ru1−N6, 171.03(8)°; N5−Ru1−N10, 167.78(8)°; N9−Ru1−N2, 171.67(8)°. For mer-[Ru(Phpytri)3]2+ (Figure 3d): Ru1−N1, 2.079(4) Å; Ru1−N2, 2.014(4) Å; Ru1−N5, 2.082(4) Å; Ru1−N6, 2.032(3) Å; Ru1−N9, 2.067(4) Å; Ru1−N10, 2.035(4) Å; N1−Ru1−N2, 78.1(1)°; N5−Ru1−N6, 78.8(1)°; N9−Ru1−N10, 78.8(1)°; N1−Ru1−N9, 168.9(1)°; N5−Ru1−N10, 167.2(1)°; N2−Ru1−N6, 173.7(1)°. For fac-[Ru(hexpytri)3]2+ (Figure 3e): Ru1−N1, 2.089(3) Å; Ru1−N2, 2.029(4) Å; Ru1−N5, 2.090(4) Å; Ru1−N6, 2.040(4) Å; Ru1−N9, 2.089(4) Å; Ru1−N10, 2.031(3) Å; N1−Ru1−N2, 78.5(1)°; N5−Ru1−N6, 78.4(1)°; N9−Ru1−N10, 77.9(2)°; N1−Ru1−N10, 173.2(1)°; N5−Ru1−N2, 174.1(1)°; N9−Ru1−N6, 174.7(2)°. For mer-[Ru(hexpytri)3]2+ (Figure 3f): Ru1−N1, 2.087(2) Å; Ru1−N2, 2.019(2) Å; Ru1−N5, 2.081(2) Å; Ru1−N6, 2.029(2) Å; Ru1−N9, 2.086(2) Å; Ru1−N10, 2.045(2) Å; N1−Ru1−N2, 78.44(8)°; N5− Ru1−N6, 78.55(8)°; N9−Ru1−N10, 78.38(8)°; N1−Ru1−N6, 169.04(8)°; N5−Ru1−N9, 174.46(8)°; N2−Ru1−N10, 174.13(8)°.

Electrochemistry. An electrochemical study of the cationic ruthenium(II) R-pytri complexes [Ru(R-pytri)3](PF6)2 was undertaken using cyclic and differential pulse voltammetry in DMF solution. A representative voltammogram is illustrated in

previously in other R-pytri complexes and has been attributed to a decrease in the π-donors d-orbital energy level, because the 1,2,3-triazole unit is a better π-acceptor than the pyridine, which it replaces.12b,25,28 D

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comprehensively explored.24,25 For our series of [Ru(Rpytri)3](PF6)2 complexes, cathodic scans show three sequential ligand-based reductions (Figure 5). The first is quasireversible and occurs at E° = −1.68, −1.64, or −1.7 V for benzyl-, phenyl-, and alkyl-substituted complexes, respectively, consistent with their inductive properties [E° = (Epc + Epa)/2]. No variation is observed between mer and fac isomers, and E° is independent of alkyl tail length. The second quasi-reversible reduction at ca. − 2 V shows a significantly larger shift between phenyl and benzyl/alkyl variants, but, more importantly, E° for the mer isomer is uniformly observed at a potential 30−40 mV more negative than its fac counterpart. The origin of this subtle difference is not clear. A third irreversible reduction occurs with Epc > −2.2 V. Stability Experiments. Since previous work had shown that related iron(II) R-pytri complexes were unstable22 in dimethyl sulfoxide (DMSO)29 and in the presence of biological ligands such as histidine, we examined the stability of some of the [Ru(R-pytri)3](PF6)2 complexes using 1H NMR competition experiments. Pleasingly, the mer- and fac-[Ru(octpytri)3](PF6)2 isomers proved stable when dissolved in d6-DMSO at room temperature (298 K). Furthermore, 1H NMR experiments in d6-DMSO at 50 °C indicated that both the mer- and fac-[Ru(octpytri)3](PF6)2 isomers were stable over a period of three days. No significant changes in the spectrum were observed and no signals due to “free” octpytri ligand could be detected. The stability of the mer- and fac-[Ru(octpytri)3](PF6)2 isomers in the presence of histidine, a common biological ligand, was then examined. One of the ruthenium complexes, either mer- or fac-[Ru(octpytri)3](PF6)2 (1 equiv) and DLhistidine (6 equiv) were dissolved in d6-DMSO and the solution was heated at 50 °C for a period of 2 days. The mixture was analyzed by 1H NMR spectroscopy at specific intervals of time over the 2 days (see Section 8 (Figure S47) in the Supporting Information). Similar to what was observed in neat d6-DMSO solution, there was no indication of ruthenium complex degradation in the collected 1H NMR spectra of the mixtures. No new signals due to degradation products or free ligand were detected. After the completion of both competition experiments, the samples were analyzed by HR-ESI-MS. The mass spectra of the mixtures only showed two major peaks, consistent with the [Ru(octpytri) 3 ] 2 + and [Ru(octpytri)3(PF6−)]+ ions; no adducts involving histidine were observed. This behavior is similar to that observed for a related [Ru2(dipytri)3]4+ helicate (where dipytri = di(2-(1-R-1H-1,2,3triazol-4-yl)pyridine) ligand linked by a 1,4-xylene spacer unit) and confirmed that the ruthenium(II) R-pytri complexes are considerably more kinetically robust than the iron(II) analogues.22 Antimicrobial Activity. Having confirmed that the ruthenium(II) R-pytri complexes, [Ru(R-pytri)3](PF6)2, displayed good stability in the presence of the biological nucleophile histidine and DMSO, we evaluated their antibacterial activity against S. aureus (ATCC 25923) and E. coli (ATCC 25922) using disk diffusion, disk dilution, and broth microdilution methods (see Table 1). Because of the lack of aqueous solubility of the compounds, DMSO stock solutions were used for the testing, and control experiments showed that DMSO had no discernible biological effect (Table 1). We examined the series of R-pytri ligands and complexes for activity (Table 1 and Section 9 in the Supporting Information). While none of the ligands displayed activity against E. coli,

Figure 4. Selected UV-vis spectra (CH3CN, 1 × 10−5 M) of the mer[Ru(R-pytri)3](PF6)2 complexes and the related [Ru(octpytri)(bipy)2](PF6)2 complex.

Figure 5, and numerical data are presented in Table 1 and Section 7 (Table S.1) in the Supporting Information. For the

Figure 5. Cyclic voltammogram of mer-[Ru(Bnpytri)3]2+, 100 mV s−1, in 0.1 M Bu4NPF6/DMF vs [Fc*]+/0 = 0.00 V.

complexes [Ru(R-pytri)3](PF6)2, anodic scans show a quasireversible oxidation associated with the RuIII/II couple. For alkyl-substituted triazoles, this occurs at E° = 1.35 V and no variation with increasing chain length was observed. For the [Ru(Phpytri)3]2+ complex, the combined effect of three electron-withdrawing phenyl triazole substituents was a ca. 70 mV anodic shift of E°(RuIII/II) to higher potential. This behavior is consistent with that observed previously by Fletcher and co-workers.24 There is no variation of the RuIII/II potential (within the experimental uncertainty) between mer- and fac[Ru(R-pytri)3](PF6)2 isomers (see Table 1). The electrochemistry of the Bnpytri and Phpytri ligands in DMF has been reported.11d An irreversible reduction is observed with Epc ≈ − 2.3 V. Withdrawal of electron density upon coordination to a metal predictably shifts the ligand reduction to more anodic potential. In each case for Pd and Pt,11d,12b and Re,12b,c the ligand-based reduction remains irreversible. Several Ru(II) complexes that incorporate a single R-pytri (with bpy or similar ligands) have been prepared.11a,12d,25,27d In each case, the R-pytri-based reduction is irreversible12b or is not reported. Similarly, the reduction electrochemistry of the [Ru(R-pytri)3]2+ systems has not been E

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Table 1. Zone of Inhibition in mm and Minimum Inhibitory Concentrations (MIC, μg/mL) for the [Ru(R-pytri)3](PF6)2 Complexes against S. aureus and E. coli Bacteria (Electrochemical Data for the Oxidation of the [Ru(R-pytri)3](PF6)2 Complexes Are Also Shown) S. aureus (ATCC 25923)

a

E. coli (ATCC 25922)

compound

zone of inhibition (mm)

MIC (μg/mL)

zone of inhibition (mm)

MIC (μg/mL)

E°(ox) for RuIII/II (V)

fac-[Ru(Bnpytri)3](PF6)2 mer-[Ru(Bnpytri)3](PF6)2 fac-[Ru(Phpytri)3](PF6)2 mer-[Ru(Phpytri)3](PF6)2 fac-[Ru(butpytri)3](PF6)2 mer-[Ru(butpytri)3](PF6)2 fac-[Ru(hexpytri)3](PF6)2 mer-[Ru(hexpytri)3](PF6)2 fac-[Ru(octpytri)3](PF6)2 mer-[Ru(octpytri)3](PF6)2 fac-[Ru(dodecpytri)3](PF6)2 mer-[Ru(dodecpytri)3](PF6)2 fac-[Ru(hexdecpytri)3](PF6)2 mer-[Ru(hexdecpytri)3](PF6)2 fac-[Ru(octpytri)(bipy)2](PF6)2 [Ru(phen)2(Me2bipy)](PF6)2 [Ru(Me4phen)3](PF6)2a cis-[Ru(bipy)2Cl2] gentamicin DMSO

11 11 12 12 10 10 19 21 11 13 10 10 10 10 11 nil b nil 26 nil

128 128 128 128 128 128 8 8 4 4 128 128 128 128 256 − 0.5 −