Fullerenes have been extensively studied for their possible applications in biological, medical, and environmental settings. Recent evidence has shown that fullerene aggregates suspended in water exert a toxic effect on eukaryotic and prokaryotic cells alike. Here we aimed to replicate the antibacterial effect of a fullerene water suspension on Escherichia coli B23, and further intended to determine whether the toxic effect of the fullerene suspension was due to compromised integrity of cellular membrane structures. We utilized three different experiments to evaluate this hypothesis: an assay for growth and viability, a fluorescent staining assay, and an intracellular pH assay. The growth and viability assay confirmed results from previous studies that fullerene aggregates indeed have an inhibitory effect on E. coli B23. However, the fluorescent staining demonstrated that exposure to the fullerene water suspension does not disrupt the structure of cellular membranes. The internal pH assay was inconclusive. Taken as a whole, our results suggest that the antibacterial effect of fullerene on E. coli B23 is not due to damage to the cellular membrane structures, but to other mechanisms that warrant further investigation. The Nobel Prize-winning discovery of buckminsterfullerene and its unique properties in 1985 (19) combined with advances in mass production techniques of the spherical C 60 molecule (26) have launched a massive wave of research into the possible applications of this novel nanoparticle. Numerous biological and medical applications have been proposed for fullerenes and its modified derivatives, ranging from anti-HIV and antimicrobial therapies to drug delivery and diagnostic techniques (2, 6). Deleterious effects of fullerene on biological systems have also been documented: it has been shown to have antibacterial properties (21) and cause oxidative stress in several fish species Computer simulation studies demonstrated that a fullerene water suspension, herein referred to as nC 60 , may be able to bind and denature nucleotides (34). Based on these findings, previous studies aimed to elucidate if fullerene exerts its toxic effects by interacting with and altering bacterial DNA Evidence provided in a separate computer simulation study by Chang and Violi showed that hydrophobic nC 60 particles could penetrate and remain within the hydrophobic center of lipid bilayers (9). The presence of the round nC 60 particle within the bilayer was shown to distort and alter the packing of membrane lipids, ultimately leading to the formation of pores within the membrane (9). However, these results were purely theoretical and the study provided no in vivo confirmation of a disruptive effect of nC 60 on biological membranes. In this study we aimed to demonstrate that nC 60 exerts an antimicrobial effect on Escherichia coli B23 in vivo, and that this effect is due to disruption of the bacterial cell membrane by nC 60 particles. While our results indicate that incubation with nC 60 inhibited the growth and viability of E. coli B23, the mechanism of inhibition does not appear to be due to disruption of the cell membrane. MATERIALS AND METHODS Bacterial strain and culture conditions. Wild type Escherichia coli strain B23 was obtained from the MICB 421 culture collection at the University of British Columbia Department of Microbiology and Immunology. Overnight cultures were grown in M9 minimal salts media (35) (supplemented with 0.2% w/v glycerol) with a loopful of E. coli B23 and incubated at 37°C in an aerator shaking at 200 rpm.

ii Nanotechnology has been undergoing tremendous development in recent decades, driven by realized perceived applications of nanomaterials in electronics, therapeutics, imaging, sensing, environmental remediation, and consumer products. Along with these developments there have been increased evidences that engineered nanomaterials are often associated with hazardous effects they invoke in biological and ecosystems through intentional designs or unintentional discharge. Consequently there is a crucial need for documenting and understanding the interactions between nanoparticles and biological and ecosystems. This dissertation is aimed at bridging such a knowledge gap by examining the biological and ecological responses to carbon nanoparticles, a major class of nanomaterials which have been mass produced and extensively studied for their rich physical properties and commercial values. Chapter I of this dissertation offers a comprehensive review on the structures, properties, applications, and implications of carbon nanomaterials, especially related to

There is an urgent need to develop efficient and rapid strategies in order to characterize the potential health risks associated with nanomaterials, given the speed with which applications and uses are increasing. Use of standard toxicity methods will not be sufficient to meet this need. This article proposes the adoption of two novel guidances: the system's biological approach to toxicity testing advocated by the US National Research Council and a nanobiological perspective that identifies key events at the nanoscale that are relevant to signal transduction and structural biology.

Abstract With the growing use of nanomaterials in bioapplications, the nanotoxicity of new nanomaterials has become a safety concern when used in various applications. In this chapter, technical developments on carbon nanotubes are described including a historical account, experimental models and potential bioapplications. Carbon nanotube (CNT) materials display superior properties in electric current carrying capacity, thermal conductivity, and thermal stability. Due to the unique CNT structure with high-aspect ratio, CNT may show unusual toxicity and complicate its safe use in a target tissue. To test nanotoxicity of CNT, we describe a set of protocols of prior knowledgebased physical and chemical characteristics to develop 3-dimensional in vitro models of the intact skin, and a 3D in vitro model of the human airway using a co-culture of normal human bronchial epithelial cells and normal human fi broblasts. The human airway 3D model served as a tool of health risk assessment of CNTs on the human respiratory systems. To test functionality at different CNT concentrations in a 3D model, physical characteristics of multiwalled CNTs and production of nitric oxide (NO) served as cell viability and infl ammatory marker; mitochondrial activity (MTT assay) served as the cytotoxic response of the epithelial cell layers; transepithelial electrical resistance (TER) measured nanotoxicity in the changes in airway physiological function. Cytoxicity and infl ammatory responses of CNTs were dependent on different size, mass, shape, and functionality of CNTs as viable in vivo tests were conducted to evaluate the

A nanobiological approach to nanotoxicology