The role of capsular extracellular polymeric substances (EPS) at the surface of planktonic microorganisms was investigated for possible toxicity mitigation from titanium dioxide (TiO₂) nanoparticles, using variable EPS producing wild-type and isogenic mutant strains of Pseudomonas aeruginosa. Membrane integrity assays revealed that increased capsular EPS reduced cell membrane damage. Acting as a barrier to the cell membrane, capsular EPS permitted attachment of nanoparticles to the cell, while simultaneously delaying cellular damage caused by the production of reactive oxygen species (ROS). Modulations in ROS production were monitored in situ; while changes in the chemical composition of the microorganisms before and after exposure were examined with Fourier transform infrared spectroscopy (FTIR). The addition of methanol, a known radical scavenger, was shown to vastly reduce ROS production and membrane integrity losses, while not affecting physical interactions of nanoparticles with the microorganism. The results support that EPS provides an attachment site for nanoparticles, but more importantly act as a barrier to cell membrane oxidation from ROS. These observations provide better understanding of the overall importance of ROS in TiO₂ microbial toxicity.