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Posted: May 19, 2008
The role of surfactants in carbon nanotube toxicity
(Nanowerk Spotlight) Synthesized carbon nanotubes, especially single-walled carbon nanotubes (SWCNTs), are in the form of bundles with other impurities such as catalyst particles and amorphous carbon debris. In order to be useful for many types of applications, for instance in nanoelectronic devices or biomedical applications, SWCNTs need to be purified and dispersed into individual nanotubes. One method to do this is by surfactant stabilization of the hydrophobic nanotube surface, which overcomes the van der Waals forces among the nanotubes and results in suspensions of individual SWCNTs.
Several commercial surfactants, such as sodium dodecyl sulfate (SDS), sodium cholate (SC) and sodium dodecylbenzene sulfonate (SDBS) have been demonstrated to efficiently disperse bundled single-walled carbon nanotubes into aqueous suspensions of individual nanotubes. However, there has been no systematic study on the toxicity of nanotube conjugates with the extensively-used SDS, SDBS, and SC surfactants. It is critical to understand the cytotoxicity of the nanotube-surfactant conjugates since these reagents are increasingly being used in industry and laboratories. The toxicity of a surfactant may not only cause health issues for those working with carbon nanotubes, but may prove toxic to cells when used as non-viral transporters for biomolecules including DNA, siRNA and proteins for therapeutic purposes.
Researchers have now investigated the cytotoxicity of SWCNTs suspended in various surfactants. Their experimental results show that the conjugates SDS/CNT and SDBS/CNT are toxic to astrocytoma cells due solely to the toxicity of the SDS and SDBS molecules, which administered alone are toxic to the cells even at a low concentration of 0.05 mg per ml within 30 min. However, the proliferation and viability of the astrocytoma cells were not affected by SWCNTs and the conjugates SC/CNT and DNA/CNT.
"Our data suggest that the cytotoxicity of the surfactant needs to be considered when selecting a surfactant for the dispersion of SWCNTs" Dr. Lifeng Dong tells Nanowerk. "The toxicity of a surfactant may not only cause health issues for those working with SWCNTs, but may prove toxic to cells when used as non-viral transporters for biomolecules. Since similar issues exist for the surfactants used to disperse other nanoscale materials, the approach outlined in our study can be used to evaluate the cytotoxicity of other nanoscale materials as well."
Dong, an Assistant Professor in the Department of Physics, Astronomy, and Materials Science at Missouri State University (MSU), together with his collaborators, investigated the cytotoxicity caused by surfactant stabilization of SWCNTs by phase contrast light microscopy characterization in combination with an absorbance spectroscopy cytotoxicity analysis. The findings were reported in the May 15, 2008 online edition of Nanotechnology ("Cytotoxicity of single-walled carbon nanotubes suspended in various surfactants"). This work has been an interdisciplinary research effort by Dong together with Prof. Colette M. Witkowski and Prof. Michael M. Craig of the Department of Biomedical Sciences at MSU.
"Our experiments demonstrate that the cytotoxicity of the nanotube-surfactant conjugates was controlled by the surfactant molecules attached on the nanotube surface" Dong explains. "Nanotubes alone do not affect the proliferation of the cells. We used a CellTiter 96 Aqueous One Solution (Promega) assay to quantitatively investigate cell proliferation and obtained assay data consistent with morphological observations obtained by phase contrast microscopy. This means that the CellTiter 96 assay components do not react with carbon nanotubes and therefore can be used to study the toxicity of carbon nanotubes and their conjugates with other functional groups on their surfaces."
Currently, there are conflicting conclusions concerning the toxicity of carbon nanotubes for both in vivo and in vitro experiments (see our Spotlight: Comparing apples with oranges – the problem of nanotubes risk assessment). Some experiments indicate that introduction of carbon nanotubes into growth medium does not affect cell proliferation and viability. However, other experiments demonstrate that structural variants of carbon nanotubes affect cell proliferation.
In Dong's study, the nanotubes were purified without metallic catalyst particles and amorphous carbons. Both catalyst particles and amorphous carbon have been demonstrated to influence cell viability. "Our experiments show that the toxicity of nanotube conjugates appears to be controlled by the specific molecules attached on the nanotube surfaces" says Dong. "This means, one criteria whether nanotubes are toxic or not depends on how the nanotubes are purified and dispersed as well as the surface cargos that they may carry. Our experiments also demonstrate that, in contrast to SDS and SDBS, SC is an environmental-friendly reagent to be used for dispersion and purification of nanotubes.
Dong and his collaborators are already taking the next steps in their research. They are now planning to utilize the methodology employed in this paper to study the toxicity of carbon nanotube conjugates for other eukaryotic and prokaryotic cells. They are also planning to study how nanotubes gain entry into cells.
Dong points out that the lack of understanding about the cytotoxicity mechanisms of carbon nanotubes is related to a lack of understanding about cell entry mechanisms. "The translocation pathway of carbon nanotubes across cell membranes remains to be elucidated. Some experiments show that nanotubes behave like cell penetrating peptides. However, other experiments reveal that cell entry of carbon nanotubes depends on temperature and can be influenced by endocytosis inhibitors."