Current high-throughput approaches evaluating toxicity of chemical agents toward bacteria typically rely on optical assays, such as luminescence and absorbance, to probe the viability of the bacteria. However, when applied to toxicity induced by nanomaterials, scattering and absorbance from the nanomaterials act as interferences that complicate quantitative analysis. Herein, we describe a bacterial viability assay that is free of optical interference from nanomaterials and can be performed in a high-throughput format on 96-well plates. In this assay, bacteria were exposed to various materials and then diluted by a large factor into fresh growth medium. The large dilution ensured minimal optical interference from the nanomaterial when reading optical density, and the residue left from the exposure mixture after dilution was confirmed not to impact the bacterial growth profile. The fractions of viable cells after exposure were allowed to grow in fresh medium to generate measurable growth curves. Bacterial viability was then quantitatively correlated to the delay of bacterial growth compared to a reference regarded as 100% viable cells; data analysis was inspired by that in quantitative polymerase chain reactions, where the delay in the amplification curve is correlated to the starting amount of the template nucleic acid. Fast and robust data analysis was achieved by developing computer algorithms carried out using R. This method was tested on four bacterial strains, including both Gram-negative and Gram-positive bacteria, showing great potential for application to all culturable bacterial strains. With the increasing diversity of engineered nanomaterials being considered for large-scale use, this high-throughput screening method will facilitate rapid screening of nanomaterial toxicity and thus inform the risk assessment of nanoparticles in a timely fashion.

Lithium intercalation compounds such as nickel manganese cobalt oxides (Li x Ni y Mn z Co1–y–z O2, 0 < x, y, z < 1, or NMCs) are complex transition metal oxides of increasing interest in nanoscale form for applications in electrochemical energy storage and as tunable catalysts. These materials exhibit sheetlike structures that expose low-energy basal planes and higher-energy edge planes in relative amounts that vary with the nanoparticle morphology. Yet there is little understanding of how differences in nanoparticle morphology and exposed crystal planes affect the biological impact of this class of technologically relevant nanomaterials. We investigated how changing nanoparticle morphology from two-dimensional (001)-oriented nanosheets to three-dimensional nanoblocks affects the release of ions and the resulting biological impact using Shewanella oneidensis MR-1 as a model organism. NMC nanoparticles were synthesized in sheetlike morphology and then converted to block morphologies by heating, leading to two morphologies of identical chemical composition that were compared to a commercially available NMC. Ion dissolution studies reveal that NMC nanomaterials release transition metal ions incongruently (Ni > Co > Mn) in amounts that vary with nanoparticle morphology. However, when normalized by the specific surface areas, the rates of release of each transition metal from flakes, blocks, and commercial material were equivalent. Similarly, the impact on S. oneidensis MR-1 was different when using mass-based dosing but was nearly identical using surface-area-normalized exposure dosing. Our results show that even though nanosheets and nanoblocks expose different crystal faces with significantly different surface energies, the rate of ion release is independent of the distribution of crystal faces exposed and depends only on the total surface area exposed. These data suggest that the key protonation steps that control release of transition metals do not depend on the degree of coordination of the initially exposed surface, providing insights into the molecular-level factors that influence the environmental impact of complex metal oxide nanomaterials. Our results have significant implications for establishing methodologies to assess toxicity of reactive nanomaterials.

Nickel-enriched lithium nickel manganese cobalt oxide, an increasingly used complex metal oxide, has unexpected dissolution behavior and impacts on two model environmental organisms.