Design and engineering of graphene-based functional nanomaterials for effective antimicrobial applications has been attracting extensive interest. In the present study, graphene oxide quantum dots (GOQDs) were prepared by chemical exfoliation of carbon fibers and exhibited apparent antimicrobial activity. Transmission electron microscopic measurements showed that the lateral length ranged from a few tens to a few hundred nanometers. Upon reduction by sodium borohydride, whereas the UV-vis absorption profile remained largely unchanged, steady-state photoluminescence measurements exhibited a marked blue-shift and increase in intensity of the emission, due to (partial) removal of phenanthroline-like structural defects within the carbon skeletons. Consistent results were obtained in Raman and time-resolved photoluminescence measurements. Interestingly, the samples exhibited apparent, but clearly different, antimicrobial activity against Staphylococcus epidermidis cells. In the dark and under photoirradiation (400 nm), the as-produced GOQDs exhibited markedly higher cytotoxicity than the chemically reduced counterparts, likely because of (i) effective removal by NaBH₄ reduction of redox-active phenanthroline-like moieties that interacted with the electron-transport chain of the bacterial cells, and (ii) diminished production of hydroxyl radicals that were potent bactericidal agents after chemical reduction as a result of increased conjugation within the carbon skeletons.