Cell-cell separation of a polychlorinated biphenyl (PCB)-degrading bacterium Comamonas testosteroni TK102 was monitored by flow cytometry. When monohydroxy metabolites of biphenyl (BP) (2-hydroxybiphenyl and 3-hydroxybiphenyl) were added to the culture, cell-cell separation of strain TK102 was inhibited at stationary phase. This inhibition was reproduced on non- PCB degrading bacteria such as Pseudomonas putida PpY101 and Escherichia coli MV1184, but was not observed on Pseudomonas aeruginosa PAO1. An opportunistic pathogen, P. aeruginosa PAO1, produces exopolysaccharide, which is known to scavenge damaging chemicals such as reactive oxygen species (ROS). The higher level of ROS and lipid peroxidants were detected in the cells treated by monohydroxybiphenyls. Fat-soluble vitamin E, which is a lipid radical scavenger, maintained bacterial cell separation during monohydroxybiphenyls treatment. Our results demonstrated that intracellular oxidative stress played an important role in the inhibition of bacterial cell separation during BP metabolism. This study shows that metabolites of environmental pollutants, such as monohydroxylated BP, inhibit bacterial cell separation by oxidative stress.