A series of fluoro-quinolones was evaluated for antibacterial activity and DNA-gyrase inhibitory potency. The relative enhancement of the overall antibacterial activity resulting from C-6 or C-8 fluorine substituent was not accompanied with a major change of DNA-gyrase inhibitory potency. Since the discovery of nalidixic acid by LESHER in 1962*, the quinolone antibacterials have emerged as a significant class of chemotherapeutic agents. The major structural change in structure-activity of both quinolone and naphtyridone series was the introduction of a fluorine at C-6. This gave rise to compounds which were dramatically more potent than nalidixic acid in vitro1-3. This modification continues to be a structural feature of all current synthetic analogues and is one of the major factors for the greatly increased activity of all of the current quinolones. However, the precise role of C-6 fluorine substituent has never been clarified. As a means of gaining additional information on the role displayed by different substitutions of fluorine atom(s) at several positions of quinolones, our group has recently shown that C-6 fluoro atom can be moved to the C-8 position while incurring only minor loss of potency4. The primary mechanism of action of quinolones is inhibition of DNA-gyrase activity5. The aim of this report was to determine the role displayed by fluorine atom(s) at C-5, C-6 and C-8 of a series of 7-(3(S)aminopyrrolidinyl)-1-cyclopropyl-1,4-dihydro-4-oxo-3-quinoline carboxylic acids at the cellular and molecular level. We compared the antimicrobial activity and inhibitory potency of these compounds against E. coli DNA-gyrase.