Comparative eco-toxicity of nanoscale TiO₂, SiO₂, and ZnO water suspensions

Abstract

The potential eco-toxicity of nanosized titanium dioxide (TiO₂), silicon dioxide (SiO₂), and zinc oxide (ZnO) water suspensions was investigated using Gram-positive Bacillus subtilis and Gram-negative Escherichia coli as test organisms. These three photosensitive nanomaterials were harmful to varying degrees, with antibacterial activity increasing with particle concentration. Antibacterial activity generally increased from SiO₂ to TiO₂ to ZnO, and B. subtilis was most susceptible to their effects. Advertised nanoparticle size did not correspond to true particle size. Apparently, aggregation produced similarly sized particles that had similar antibacterial activity at a given concentration. The presence of light was a significant factor under most conditions tested, presumably due to its role in promoting generation of reactive oxygen species (ROS). However, bacterial growth inhibition was also observed under dark conditions, indicating that undetermined mechanisms additional to photocatalytic ROS production were responsible for toxicity. These results highlight the need for caution during the use and disposal of such manufactured nanomaterials to prevent unintended environmental impacts, as well as the importance of further research on the mechanisms and factors that increase toxicity to enhance risk management.

Introduction

Titanium dioxide (TiO₂), silicon dioxide (SiO₂), and zinc oxide (ZnO) are common additives with a variety of applications. TiO₂ is a good opacifier and is used as a pigment in paints, paper, inks, and plastics. Crystalline SiO₂ is employed in electronics manufacturing as both semiconductor and electrical insulator. The ceramic nature of ZnO permits its function as both pigment and semiconductor. Nanoscale TiO₂, SiO₂, and ZnO offer greater surface area than their bulk counterparts, allowing for improved performance in established applications.

Accompanying the well-established use of TiO₂, SiO₂, and ZnO, research has been conducted on their potential toxicity (Rincon and Pulgarin, 2004; Lonnen et al., 2005). A wealth of information exists on the toxicity of TiO₂ towards bacteria (e.g. Wei et al., 1994; Block et al., 1997; Kwak et al., 2001). TiO₂ is reputed to be toxic to both Gram-negative and Gram-positive bacteria. In a mixed culture experiment, an unidentified Gram-positive Bacillus subtilis was less sensitive than a pure culture of Gram-negative Escherichia coli to the effects of TiO₂, possibly due to the ability of B. subtilis to form spores (Rincon and Pulgarin, 2005). However, other studies have found Gram-positive bacteria to be more sensitive than Gram-negative bacteria to the antibacterial effects of TiO₂ (Fu et al., 2005). The antibacterial properties of TiO₂ have been exploited in water treatment reactors. A concentration of TiO₂ ranging from 100 to 1000 ppm has been reported to completely disinfect water containing 10⁵–10⁶ E. coli cells per ml in 30 min under illuminated conditions (Wei et al., 1994; Maness et al., 1999).

Fewer studies have been initiated on the antibacterial activities of either SiO₂ or ZnO. Bulk SiO₂ has been used as a control particle in several studies due to its postulated lack of toxicity towards bacteria (e.g. Liang et al., 2004). ZnO has been reported to exhibit antibacterial activity with Gram-positive B. subtilis being more sensitive to its effects than the Gram-negative E. coli (Sawai et al., 1995). The minimal inhibitory concentrations ranged from 2000 to 12,500 ppm for B. subtilis and 50,000 to 100,000 ppm for E. coli depending on particle size (Sawai et al., 1996). While these data suggest that ZnO is much less toxic to E. coli than TiO₂, it is not possible to directly compare these studies due to differences in experimental design (e.g., particle size, concentration of bacteria, application of light).

The differential toxicity of TiO₂, SiO₂, and ZnO may be related to the mechanisms by which the particles act on cells. It is documented that these three compounds are photosensitive and produce reactive oxygen species (ROS) in the presence of light (Yeber et al., 2000; Fubini and Hubbard, 2003; Kubo et al., 2005). However, a positive correlation between photocatalytic ROS production and antibacterial activity has been determined only for TiO₂. Light in these reactions is usually provided by specific wavelength high-intensity lamps; however, one study showed that TiO₂ exhibited antibacterial properties when sunlight was the source of illumination (Wei et al., 1994).

In previous studies, TiO₂ particles that were toxic to bacteria ranged in size from tens of nanometers to hundreds of micrometers. It is not currently clear whether particle size is a key determinant of toxicity or whether surface chemistry and morphology are more important. With the rapid emergence of nanoparticles, it is important to identify the factors that accentuate toxicity. Currently, legislation of nanomaterials is limited, mainly due to the lack of toxicological information and the novelty of the field (Hogue, 2005). However, it is crucial that we understand the fate and impact of potential “contaminants” to permit the development of appropriate disposal mechanisms that mitigate the contamination of surface and groundwater resources.

Little published research has focused on the antibacterial effects related to disposal or accidental spillage of TiO₂, SiO₂, and ZnO. Many studies using nanoscale TiO₂ have incorporated solublising agents (e.g., hydroxyl groups) into the suspension (Kwak et al., 2001) or have immobilised the TiO₂ onto glass (Rincon and Pulgarin, 2004), stainless steel (Yu et al., 2003) or acetate sheets (Lonnen et al., 2005) or have utilized artificial (relatively intense) light sources. While these studies focused on parameters of their particular application, they might not be representative of the effect of raw nanoscale TiO₂ release into the aqueous environment. Therefore, we used nanoparticle water suspensions and natural sunlight to better model natural nanoparticle exposure.

This paper compares and contrasts the toxic effects associated with TiO₂, SiO₂, and ZnO water suspensions using two model bacterial species, Gram-negative E. coli and Gram-positive B. subtilis. The objectives of this study were to (a) determine the concentrations at which the three suspensions are toxic to our test organisms, (b) determine whether the size of the released nanoparticle affects antibacterial activity, and (c) determine whether natural light stimulates toxicity of the nanoparticles to bacteria.