Fluoroquinolone-derived fluorescent probes for studies of bacterial penetration and efflux

Abstract

Fluorescent probes derived from the fluoroquinolone antibiotic ciprofloxacin were synthesised using a Cu(i)-catalysed azide-alkyne cycloaddition (CuAAC) to link a ciprofloxacin azide derivative with alkyne-substituted green and blue fluorophores. The azide (2) and fluorophore (3 and 4) derivatives retained antimicrobial activity against Gram-positive and Gram-negative bacteria. The use of confocal fluorescent microscopy showed intracellular penetration, which was substantially enhanced in the presence of carbonyl cyanide 3-chlorophenylhydrazone as an efflux pump inhibitor in Escherichia coli.

Scheme 1. Reagents and conditions: (i) NaHCO₃, NaI, acetonitrile, reflux 12–24 h, quantitative; (ii) CuSO₄, sodium ascorbate, DMF/H₂O, 57%; (iii) CuSO₄, sodium ascorbate, DMF/H₂O, 12%.

Fig. 1. Confocal fluorescent microscopy of live S. aureus labelled with A: red FM4-64FX membrane dye, green cipro-NBD 3, blue nucleic acid dye Hoescht 33342, and overlay; B: red FM4-64FX membrane dye, blue cipro-DMACA 4, green nucleic acid dye Syto-9, and overlay.

Fig. 2. Confocal fluorescent microscopy of: live E. coli AG102 labelled with: red FM4-64FX membrane dye and green cipro-NBD probe 3, with (A, several fields of view showing internalisation) and without (B) efflux pump inhibitor, showing no internalisation.