Computer simulation study of fullerene translocation through lipid membranes JIRASAK WONG-EKKABUT 1,2† , SVETLANA BAOUKINA 1 , WANNAPONG TRIAMPO 2,3 , I-MING TANG 2,4 , D. PETER TIELEMAN 1 AND LUCA MONTICELLI 1 * ‡ 1 Department of Biological Sciences, University of Calgary, Calgary, Alberta T2N 1N4, Canada 2 Department of Physics, Faculty of Science, Mahidol University, Bangkok 10400, Thailand 3 Center of Excellence for Vectors and Vector-Borne Diseases, Mahidol University at Salaya, Nakhonpathom 73170, Thailand 4 Center of Nanoscience and Nanotechnology, Mahidol University, Bangkok 10400, Thailand † Present address: Department of Applied Mathematics, University of Western Ontario, London, Ontario, Canada ‡ Present address: Laboratory of Physics, Helsinki University of Technology, Espoo, Finland *e-mail: luca.monticelli@gmail.com Published online: 18 May 2008; doi:10.1038/nnano.2008.130 Recent toxicology studies suggest that nanosized aggregates of fullerene molecules can enter cells and alter their functions, and also cross the blood–brain barrier. However, the mechanisms by which fullerenes penetrate and disrupt cell membranes are still poorly understood. Here we use computer simulations to explore the translocation of fullerene clusters through a model lipid membrane and the effect of high fullerene concentrations on membrane properties. The fullerene molecules rapidly aggregate in water but disaggregate after entering the membrane interior. The permeation of a solid-like fullerene aggregate into the lipid bilayer is thermodynamically favoured and occurs on the microsecond timescale. High concentrations of fullerene induce changes in the structural and elastic properties of the lipid bilayer, but these are not large enough to mechanically damage the membrane. Our results suggest that mechanical damage is an unlikely mechanism for membrane disruption and fullerene toxicity. The extent of the production and use of nanomaterials is rapidly growing. Carbon nanomaterials, such as fullerenes and nanotubes, are among the most extensively studied nanomaterials. Bulk fullerene production at a scale of tons is already under way 1 . Although the effects of nanoparticles on health and the environment are becoming more of a concern, studies on toxicology and the environmental impact of nanoparticles are still scarce 2 . Inhaled ultrafine carbon particles deposit in the lung 3 and translocate into the brain, especially into the olfactory bulb, by means of the olfactory nerves and the blood 4,5 . Fullerene is soluble in numerous organic solvents and it forms a stable colloidal suspension in water, also known as ‘nano-C60’ 6,7 . This aggregate has been well characterized experimentally and its size lies between tens and a few hundreds of nanometres 6–9 . Experimental results suggest that, despite their large size, fullerene aggregates can penetrate cells and cross the blood – brain barrier 4 . The mechanism of nano-C60 penetration through a lipid membrane has not yet been established. The mechanism of cell membrane disruption is also not well understood. It has been reported that the toxicity of carbon nanoparticles depends on their solubility in water; for example, the cytotoxicity of pristine fullerene is seven orders of magnitude higher than for functionalized fullerenes with high solubility 10 . It has been recently proposed that fullerene causes cell membrane leakage due to lipid peroxidation 11 . On the other hand, it has also been reported that C60 and water-soluble fullerene derivatives could be used as antioxidants against radical-initiated lipid peroxidation 12 , as well as drug carriers 13,14 . The protection from lipid peroxidation was found to be higher for pristine C60 than for water-soluble derivatives 12 . Whether the biological activity of fullerenes is desirable or not, experimental evidence published so far indicates that solubility in the membrane interior is an important determinant of biological activity 5 . In the present study we describe the thermodynamics and mechanism of the permeation of fullerene aggregates through cell membranes, based on computer simulations. Previous simulation studies of nanoparticles have investigated the insertion of individual hydrophobic nanotubes in membranes 15,16 and water transport through carbon nanotubes 17,18 . Although fullerene is known to aggregate in water, simulation studies reported so far have focused on monomeric fullerene and water-soluble derivatives, and include investigations of fullerene solvation in water 19 , the interactions between fullerene molecules in vacuum 20 and in water 21 , the translocation of monomeric C60 across a lipid bilayer 22 and the interaction between two individual C60 molecules inside a lipid bilayer 23 . Following an earlier approach 24–26 , we developed a coarse-grained (CG) model based on experimental partitioning of fullerene between polar and nonpolar phases, which is the main determinant of permeation across a lipid membrane 27 . Our work provides insight into the thermodynamics of fullerene clusters permeation through cell membranes and the effect of high concentrations of fullerene on the structural and elastic properties of a lipid bilayer, and suggests that mechanical damage is not likely to be responsible for membrane disruption and fullerene toxicity. MECHANISM AND ENERGETICS OF FULLERENE PERMEATION Fullerene is not soluble in water and only marginally soluble in polar organic solvents, but its solubility in hydrocarbons is ARTICLES nature nanotechnology | VOL 3 | JUNE 2008 | www.nature.com/naturenanotechnology 363 © 2008 Nature Publishing Group