Structure-Property-Toxicity Relationships of Graphene Oxide: Role of Surface Chemistry on the Mechanisms of Interaction with Bacteria

Graphene oxide (GO) is an antimicrobial agent with tunable surface chemistry. To identify the physicochemical determinants of GO's antimicrobial activity, we generated different modified Hummer's GO materials thermally annealed at 200, 500, or 800 °C (TGO200, TGO500, and TGO800, respectively) to modify the surface oxygen groups on the material. Plating assays show that as-received GO (ARGO) and TGO200, TGO500, and TGO800 reduce Escherichia coli viability by 50% (EC₅₀) at 183, 143, 127, and 86 μg/mL, respectively, indicating higher bacterial toxicity as ARGO is reduced. To uncover the toxicity mechanism of GO, fluorescent dye-based assays were used to measure oxidative stress at the EC₅₀. ARGO showed an increase in intracellular reactive oxygen species, measured as an increase in 2′,7′-dichlorodihydrofluorescein diacetate fluorescence, whereas TGO500 and TGO800 induced an increase in the fluorescence of fluorescein diacetate (FDA) by 30 and 42%, suggesting a decrease in cell permeability. Because of a possible wrapping mechanism, plating assays after post-exposure sonication were performed to explain TGO's low oxidative response and high FDA levels. Results show no difference in colony-forming units, indicating that inhibition of cell growth is a result of the adsorption of bacterial cells on the GO material. By comparing different GO samples at their EC₅₀, this study reveals that reduction of GO alters both the mechanisms of cellular interaction and the degree of toxicity to bacteria.