Ciprofloxacin-Based Ionic Liquids Increase Mutation Frequency in Escherichia coli

Background/Objectives: Formulating antibiotics as active pharmaceutical ingredient ionic liquids (API-ILs) has been proposed as a strategy to help overcome antimicrobial resistance. However, the effects of API-ILs on bacterial mutation frequency, an increase of which is associated with a higher risk of resistance development, have not yet been assessed. Here, API-ILs based on the antibiotic ciprofloxacin were synthesized using five structurally different counter ions of varying biological activity - low ([Chol]⁺ and [EMMor]⁺), intermediate ([TMC₁₀A]⁺) and high ([TMC₁₆A]⁺ and [TC₈MA]⁺) - and investigated in terms of their antimicrobial activity and mutation frequency in Escherichia coli MG1655. Methods: API-ILs were synthesized according to the CBILS© route. Conductivities and antimicrobial activity (determined by minimal inhibitory concentrations (MICs) and disk diffusion (DD) assays) of API-ILs as well as of individual API and ILs were measured, followed by mutation frequency assays. Results: Five novel ciprofloxacin-based API-ILs were synthesized. Overall, a lower dissociation of API-ILs compared to the respective ILs was observed, indicating presence of stable ion pairs in aqueous solution. All API-ILs retained the antimicrobial activity of ciprofloxacin. A higher mutation frequency (2.6-6.99-fold increase) was observed for API-ILs than for ciprofloxacin alone (1.71-fold increase), when compared to no treatment control, while ILs alone had no or a moderate impact (0.62-1.65-fold increase). Conclusions: Although it is possible to synthesize novel stable API-IL compounds with a high antimicrobial activity using ciprofloxacin and ILs of different structural classes, this can result in increased bacterial mutation frequencies. It is therefore crucial to improve our understanding of how API-ILs can be designed in a safer way.