title: Dinuclear polypyridylruthenium(II) complexes: flow cytometry studies of their accumulation in bacteria and the effect on the bacterial membrane.
abstract: OBJECTIVE
  To determine the energy dependency of and the contribution of the membrane potential to the cellular accumulation of the dinuclear complexes [{Ru(phen)2}2{μ-bbn}](4+) (Rubbn) and the mononuclear complexes [Ru(Me4phen)3](2+) and [Ru(phen)2(bb7)](2+) in Staphylococcus aureus and Escherichia coli, and to examine their effect on the bacterial membrane.
  
  
  METHODS
  The accumulation of the ruthenium complexes in bacteria was determined using flow cytometry at a range of temperatures. The cellular accumulation of the ruthenium complexes was also determined in cells that had been incubated with the metal complexes in the presence or absence of metabolic stimulators or inhibitors and/or commercial dyes to determine the membrane potential or membrane permeability.
  
  
  RESULTS
  The accumulation of ruthenium complexes in the two bacterial strains was shown to increase with increasing incubation temperature, with the relative increase in accumulation greater with E. coli, particularly for Rubb12 and Rubb16. No decrease in accumulation was observed for Rubb12 in ATP-inhibited cells. While carbonyl cyanide m-chlorophenyl hydrazone (CCCP) did depolarize the cell membrane, no reduction in the accumulation of Rubb12 was observed; however, all ruthenium complexes, when incubated with S. aureus at concentrations twice their MIC, depolarized the membrane to a similar extent to CCCP. Except for the mononuclear complex [Ru(Me4phen)3](2+), incubation of any of the other ruthenium complexes allowed a greater quantity of the membrane-impermeable dye TO-PRO-3 to be taken up by S. aureus.
  
  
  CONCLUSIONS
  The results indicate that the potential new antimicrobial Rubbn complexes enter the cell in an energy-independent manner, depolarize the cell membrane and significantly permeabilize the cellular membrane.
authors.affiliation: School of Physical, Environmental and Mathematical Sciences, University of New South Wales, Australian Defence Force Academy, Canberra, ACT 2600, Australia.
authors.forename: Fangfei
authors.initials: F
authors.lastname: Li
authors.forename: Marshall
authors.initials: M
authors.lastname: Feterl
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Warner
authors.forename: F Richard
authors.initials: FR
authors.lastname: Keene
authors.forename: J Grant
authors.initials: JG
authors.lastname: Collins
mesh_terms: D000890:Anti-Infective Agents / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects* / Q000378:metabolism*; D005434:Flow Cytometry; D008564:Membrane Potentials / Q000187:drug effects; D017975:Ruthenium Compounds / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000378:metabolism*; D013696:Temperature
chemical_list: D000890:Anti-Infective Agents; D017975:Ruthenium Compounds
keywords: bactericidal metal complexes; cellular accumulation; membrane depolarization; membrane permeability