Cellular toxicity of titanium dioxide nanotubes and nanowires

Cellular toxicity of titanium dioxide nanotubes and nanowires

One of the complications of nanotoxicology is that the toxicity of a specific nanomaterial cannot be predicted from the toxicity of the same material in a different form. For instance, while the toxicity of inert systems such as iron oxides, gold, or silver has been investigated for nearly isotropic particles (i.e., with a low aspect ratio), the toxicity of these materials in nanofilament form cannot be predicted from their known toxicity as nanoparticles. Fully understanding the toxic mechanisms of nanoscale materials is an essential prerequisite in being able to design harmless nanomaterials whose interactions with biological cells is non-lethal.

Currently, a lot of nanotoxicological research effort is focused on carbon nanotubes, but nanofilaments are not exclusively based on carbon materials and can be produced from many inorganic materials in the form of nanotubes and nanowires. Applying the 'precautionary principle' to nanotechnology would require much more extensive nanotoxicological research on all types of nanomaterials; and there seems to be a particular lack of findings concerning non-carbon nanofilaments. Researchers in Switzerland have now taken a closer look at the fate of titanium dioxide (TiO2) based nanofilaments in the body. Their results are cause for concern.

"TiO2 nanoparticles are widely used as UV blockers in sunscreens" says Arnaud Magrez. "Their cytotoxicity has been tested before and they were found to be rather non-toxic. Our new study shows that TiO2 based nanofilaments, however, can be quite toxic. The geometry of nanoparticles appears to play a crucial role in cytotoxicity. Furthermore, the toxicity can be enhanced by the presence of defects on the nanofilament surface, resulting from chemical treatment."

These new findings clearly demonstrated that the presence of TiO2 nanofilaments (synthesized by hydrothermal treatment from anatase and highly concentrated NaOH solution) had a strong dose-dependent effect on cell proliferation and cell death. Nanofilament internalization and alterations in cell morphology were observed. Acid treatment performed to substitute Na+ with H+ in the nanofilaments strongly enhanced the cytotoxic action.

Magrez, a researcher at the NN Research Group (Laboratoire de nanostructures et nouveaux matériaux électroniques) at the Ecole Polytechnique Fédérale de Lausanne (EPFL) in Switzerland, is first author of a recent paper in ACS Nano ("Cellular Toxicity of TiO2-Based Nanofilaments"). In this work, Magrez and colleagues from his group as well as the University of Fribourg, studied the cellular toxicity of TiO2-based nanofilaments in relation to their morphology and surface chemistry.

Transmission Electron micrograph of TiO2-based nanowires. (a) Irreversible injuries and several morphological alterations of H596 epithelial cells can be observed after 2 days exposure with TiO2-based nanofibers (concentration is 2µg per ml). TiO2-based Nanofibers appear as golden wires (b).

Apart from wide-spread use of TiO2 nanoparticles, large-scale arrays of TiO2-based nanofilaments (including nanotubes and nanowires) are already being used in photovoltaic cells and in photoelectrolyzer for the production of hydrogen by water splitting.

The surface cells of the airways, including the epithelial cells of the lungs, are the first cell type to encounter TiO2-based nanofilaments released into the environment. Therefore the Swiss researchers decided to investigate the acute cytotoxicity of different TiO2-based nanofilaments on lung cells in vitro. In their experiments they evaluated the cytotoxic effect of the TiO2-based nanofilaments by the widely established MTT assay performed with H596 human lung tumor cells.

Because the MTT assay measures the combined effects of cell proliferation and metabolic activity of cells and was reported to be prone to artifacts under certain experimental conditions, the EPFL team also validated their results by directly counting the number of surviving cells from microphotographs. Magrez says that, in comparison to untreated cells, both the MTT signals and number of cells were decreased in all nanomaterial-treated samples and the two methods – MTT assay and cell counting – yielded essentially identical results.

Morphology of H596 lung carcinoma cells exposed to TiO2-based nanofilaments. Cells were treated for 4 days with 2 µg/mL TiO2 nanotubes, fixed and HE-stained. (a) Nanofilaments inside the cells have needle-like structures often concentrated around the enlarged nucleus (arrows), and even thinner nanofilaments are found within the nucleus (arrowheads). (b) In some giant cells, the nuclei are strongly lobulated or possibly fragmented, and the nanofilaments are localized between the lobules (arrows). In (a) and (b), a weaker staining of the nuclei was chosen to better visualize the intranuclear nanotubes. (c) Besides larger fibers (arrow), nanofilaments are also present in the form of small dark particles (arrowheads). Scale bar: 20 µm.

The researchers found that geometry of the nanomaterials appears to play a role, surface chemistry was the most important aspect in determining the survival of exposed cells.

"The importance of surface chemistry had already been observed in a previous report on carbon-based nanomaterials," says Magrez. "Even though multiwalled carbon nanotubes have comparable diameter and length as TiO2 nanofilaments, their toxicity is markedly different. Thus, the chemical composition of nanomaterials also appears to have an effect on cell survival. Whether the toxicity determined in this acute model will also translate to models of chronic toxicity or even tumor development (lung carcinoma or mesotheliomas) needs to be addressed in future studies."

Rewritable photonic paper

Rewritable photonic paper

Photonic crystals are a new kind of material. Also known as photonic band gap material, they are similar to semiconductors, where the electrons are replaced by photons (i.e. light). By creating periodic structures out of materials with contrast in their dielectric constants, it becomes possible to guide the flow of light through the photonic crystals in a way similar to how electrons are directed through doped regions of semiconductors. The photonic band gap (that forbids propagation of a certain frequency range of light) gives rise to distinct optical phenomena and enables one to control light with amazing facility and produce effects that are impossible with conventional optics.

Photonic crystals are very suitable for fabricating sensor devices because the optical signals of these responsive materials are tightly coupled with various external stimuli and modulations. They could also play a significant role on the way to all-optical devices in communication and information technology – they form a basis material for the future realization of optical components and circuits, and maybe even complex optical circuits or optical computers.

One application area that has seen a lot of activity recently is photonic paper and ink. One advantage of photonic papers and displays is that they are brilliant and free of glare in sunlight, which are superior to the characteristics of other emissive display technologies for outdoor usage such as advertising billboards. Another advantage is that, for recording purposes, the structural colors of photonic paper are usually more durable than traditional pigments and dyes.

However, there are still a number of issues and problems that prevent the practical applications of the photonic crystal based materials. Researchers in California have now addressed some of them by reporting a new type of rewritable photonic paper on which a durable ink mark can be written or erased by introducing or removing a hygroscopic salt in the surface layer of the paper.
The key point of their work is the use of hygroscopic salt solutions as 'ink' to swell the polymer matrix of photonic crystals and create a very durable contrast of diffraction colors on the photonic paper. The ink marks can be erased by rinsing the paper in water and drying thereafter. The rewritability of the photonic paper and low toxicity of the materials involved make this paper/ink system environmentally friendly, inexpensive, and useful for general applications involve color writing and display.

Digital photo images of flexible photonic paper (3 cm x 4 cm) on the plastic substrate with and without letters printed. The color gradient in the photos is caused by the flash lamp illuminating as a point light source.

Yadong Yin, an assistant professor in the Department of Chemistry at the University of California, Riverside, explains to Nanowerk that his lab's new invention shows several major improvements compared to existing photonic paper fabrication technologies:

1) Conventionally, the fabrication for the photonic-crystal based paper typically involves very slow processes, such as colloidal sedimentation or solvent evaporation. It usually takes days or even months to grow photonic crystals with a reasonable size, therefore limiting their practical use.

In Yin's work, the photonic paper can be produced instantaneously by assembling magnetic particles into periodic arrays using an external field. The periodic photonic structures can be immediately fixed in a polymer matrix. This allows for future mass production.

2) Conventionally, writing the color information in the photonic paper is achieved by applying a volatile solvent (ink), which swells the polymer matrix, changes the periodicity of the photonic structure, and eventually changes the color of inked region. The color disappears when the solvent evaporates. So the color writing is not permanent.

"In our work, the ink is a water-ethanol solution of inorganic hygroscopic salt" says Yin. "This salt solution does not evaporate at room temperature and keeps the polymer matrix in the swelled state so that the color information stays for a long time. Erasing of the written information can be simply achieved by removing the salt solution using additional water."

3) Unlike conventional methods which typically involve toxic organic solvent, the salt ink and rinsing water (and the photonic paper itself) are nontoxic and environmentally benign. Yin's team prepared their new type of photonic paper through the fast magnetic field-induced self-assembly of Fe3O4@SiO2 colloids, followed by a simultaneous UV curing process to fix the photonic structures inside the PEGDA matrix.

The photonic paper can be fabricated onto a glass substrate or flexible plastic substrate. The findings have been published in a paper in the July 17, 2009 online edition of Advanced Materials Schematic illustrations of the structure of the unit cell and the mechanism of writing or erasing realized by infiltrating or removing the hygroscopic salt.

Yin says that, in principle, the technique developed in his lab can potentially be used to replace conventional paper/ink systems in many areas. Also, compared to existing techniques, his team's rewritable photonic systems have several advantages including the easiness for large-area fabrication, the versatility in pattern generation, and brilliant color in bright sunlight illumination (in contrast to liquid-crystal-type displays). "We believe that more elaborate features, such as multicolor printing, may be realized in the future with the use of multiple inks and modifications to the polymeric matrix of the photonic paper"

There are still some kinks to iron out, though. For instance, the researchers experienced a problem when they write on the photonic paper manually using a small glass capillary 'pen'. As the capillary is not specially designed for writing, the delivery of 'ink' seems to be not very uniform. Yin says that he would expect a better design of a 'pen' or the integration with ink-jet printing process would solve the problem.

The immediate future work for Yin's team is to find out the ultimate resolution of this photonic paper, which is an important factor for practical applications. "One interesting question that we will keep working on is the realization of multi-color writing/printing, which might be possible if multiple ink salts are used," says Yin. "We will also explore the integration of this writing mechanism with ink-jet printing techniques in order to achieve a highly controllable delivery of inks."