Bacteria growing in vivo multiply much more slowly than in vitro. If the bactericidal activity of quinolones may be affected by an increase in generation time (g) was studied in batch cultures as well as under the well-controlled conditions of a continuous-flow culture. By limiting the nutrient supply, generation times were lengthened from approximately 0.45 to 1.5 h up to 3.9 h. Three recent clinical isolates each of Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa were exposed to twice the MIC of ciprofloxacin, norfloxacin, fleroxacin and ofloxacin. The ‘killing rates’ were calculated in analogy to the growth rate. The bactericidal activity of the quinolones tested against E. coli was minimally influenced by the reduced generation time and the effect against S.aureus was moderate. As compared to their rapidly growing counterparts (g = 0.4 h) slowly growing (g = 1.3 h) P. aeruginosa were killed even more effectively by ciprofloxacin (176% increase) fleroxacin (48% increase) norfloxacin (36% increase) and ofloxacin (86% increase). These changes may likely be due to adaptive responses of the outer membranes of the bacteria to the limited nutrient supply thereby sensitizing the bacteria to the bactericidal activity of quinolones.