likewise are employed in this role.

likewise are employed in this role.
Use in staphylococcal conjunctivitis might also be practicable. Animal experiments could be used to test these possibilities. Reasons for the lack of activity of compound 1 against any of the Gram-negative bacteria, even at 512mg/L, are uncertain but efflux wasconsideredapossiblecontributoryfactor.Insupportofthis,we have recently shown that strains of Salmonella lacking components of efflux pumps such as the pivotal resistance–nodulation–division (RND) pump AcrAB–TolC are susceptible to plant-derived antibac- terials such as falcarindiol, which is inactive against their parent strains [28]. However, the activity of compound 1 against Gram- negative isolates was not improved with the addition of the outermembranepermeabilisersethylenediaminetetra-aceticacid (1mM) or polyethyleneimine (50mg/L) (data not shown), and its molecular weight (346Da) is far below the general exclusion limit for the outer membrane. These data suggest that outer membrane impermeability is not responsible for the lack of activity. Inactivation by chemical modification is another possible mechanism for the lack of activity of compound 1 against Gram-negative bacteria, and further work will be required to investigate this possibility. Compound 1 inhibited the ATP-depended MurE ligase from M. tuberculosis, a cytoplasmic enzyme that participates in the biosynthesis of cell wall peptidoglycan [24]. The key enzymes of the cell wall peptidoglycan biosynthesis pathway are essential for the survival of many bacterial pathogens[29] and the reisrenewedinterest inthesearchfornoveltherapeutictargetsfortacklingexistingdrug resistance [30,31]. Compound 1 also had some ability to impede efflux. The low accumulation of 14C-enoxacin by SA-1199B in the absence of inhibitors was expected and is the result of overexpression of NorA in this strain (Fig. 3). Compound 1 at 50M significantly increased enoxacin accumulation and this concentration was as effective, or perhapsevenmoreeffective,than33Mreserpine.Increasedaccumulation of enoxacin is consistent with interference with NorA. As such, it is clear that compound 1 is a NorA inhibitor. Use of con- centrations that surpassed the MIC of compound 1 for SA-1199B was not problematic based on time–kill assays, which revealed no effect of compound 1 at 50M on the viability of SA-1199B over 60min. The pharmacophore of compound 1 may provide a starting point towards the development of a more effective inhibitor, and its weak cytotoxic activity towards cancer and mammalian cell lines suggests that this class of compound may have potential in a topical formulation or, if antibacterial activity in the presence of blood can be enhanced through iterative chemistry, as a lead in the development of a new class of systemic antibiotics. Compounds that are antibacterial and can inhibit efflux processes of bacteria could have potential to treat infections due to multidrug-resistant strains.
Funding:TheHeptagonFundandUnionLifeSciencesfundedthis work. Competing interests: None declared. Ethical approval: Not required.