LAUSR

While positively charged nanomaterials induce cytotoxicity in many organisms, much less is known about how the spatial distribution and presentation of molecular surface charge impacts nanoparticle-biological interactions. We systematically functionalized diamond nanoparticle surfaces with five different cationic surface molecules having different molecular structures and conformations, including four small ligands and one polymer, and we then probed the molecular-level interaction between these nanoparticles and bacterial cells. Shewanella oneidensis MR-1 was used as a model bacterial cell system to investigate how molecular length and conformation of cationic surface charges influence their interactions with the Gram-negative bacterial membranes. Nuclear magnetic resonance (NMR) and X-ray photoelectron spectroscopy (XPS) demonstrate the covalent modification of nanoparticle surface with the desired cationic organic monolayers. Surprisingly, bacterial growth-based viability (GBV) and membrane damage assays both show only minimal biological impact by the NPs functionalized with short cationic ligands within the concentration range tested. Yet NPs covalently linked to a cationic polymer induce strong cytotoxicity, including reduced cellular viability and significant membrane damage at the same concentration of cationic groups. Transmission electron mi-croscopy (TEM) images of these NP-exposed bacterial cells show that NPs functionalized with cationic polymers induce significant membrane distortion and production of outer membrane vesicles, while NPs bearing short cationic ligands exhibit only weak membrane association. Our results demonstrate that the spatial distribution of molecular charge plays a key role in controlling the interaction of cationic nanoparticles with bacterial cell membranes and subsequent biological impact. Nanoparticles functionalized with ligands having different lengths and conformations can have large differences in interactions even while having nearly identical zeta potentials. While zeta potential is a convenient and commonly used measure of nanoparticle charge, it does not capture essential differences in molecular-level nanoparticle properties that control their biological impact.