Nanoparticle dispersion in environmentally relevant culture media: a TiO₂ case study and considerations for a general approach

Nanoparticle exposure in toxicity studies requires that nanoparticles are bioavailable by remaining highly dispersed in culture media. However, reported dispersion approaches are variable, mostly study-specific, and not transferable owing to their empirical basis. Furthermore, many published approaches employ proteinaceous dispersants in rich laboratory media, both of which represent end members in environmental scenarios. Here, a systematic approach was developed to disperse initially agglomerated TiO₂ nanoparticles (Aeroxide® TiO₂ P25, Evonik, NJ; primary particle size range 6.4–73.8 nm) in oligotrophic culture medium for environmentally relevant bacterial toxicity studies. Based on understanding particle–particle interactions in aqueous media and maintaining environmental relevance, the approach involves (1) quantifying the relationship between pH and zeta potential to determine the point of zero charge of select nanoparticles in water; (2) nominating, then testing and selecting, environmentally relevant stabilizing agents; and (3) dispersing via “condition and capture” whereby stock dry powder nanoparticles are sonicated in pre-conditioned (with base, or acid, plus stabilizing agent) water, then diluted into culture media. The “condition and capture” principle is transferable to other nanoparticle and media chemistries: simultaneously, mechanically and electrostatically, nanoparticles can be dispersed with surrounding stabilizers that coat and sterically hinder reagglomeration in the culture medium.