Warheit and colleagues compared the pulmonary toxicity of different sized TiO2 particles after instillation in rat (Warheit et al, 2006). They found that "toxicity is not dependent upon particle size and surface area”. In a subsequent study with different nanoscale TiO2 test materials, they determined that crystal structure and particle surface reactivity (using the Vitamin C yellowing assay) played important mechanistic roles in facilitating adverse pulmonary effects (Warheit et al, 2007a). Determining the lung toxicity of fine- and nano-quartz in a later study they confirmed previous results (Warheit et al, 2007b). Instead of a correlation between the potency range of toxicity and particle size or surface area, they found consistency between toxicity and the results of in vitro erythrocyte haemolysis studies (as a measure of surface reactivity using a different assay). Silanol groups on the silica surface can react with carbonyl oxygen on the erythrocyte membrane. This interaction may produce ROS in amounts that exhaust the protective mechanisms of the erythrocyte and in the end lead to cell membrane rupture (Razzaboni and Bolsaitis 1990). Thus, Warheit et al, supplemented their first conclusion and claimed that "toxicity is not dependent upon particle size but on surface characteristics. However, based on a comparable surface chemistry per unit of area, the total amount of charge or reactive groups increased with decreasing particle size (or increasing surface area) but equal particulate mass.
Considering both particle size/surface area dose as well as surface characteristics, Duffin et al (2007) tested the pulmonary toxicity of different particles indicated by neutrophil content in BALF of rats after instillation. Low-toxicity, low-solubility materials (PSP, see also Section 1.3), metal nanoparticles (nickel and cobalt), and quartz as an example of a particle with a highly reactive surface were examined. PSPs revealed a greater inflammatory response than the smaller the particles (i.e. the nominally larger the surface area dose). Compared to these materials, quartz, at the same surface area dose, induced a far greater level of inflammation. The inflammatory response of the metals fell midway between the effect of the LTLS and the quartz particles Duffin et al stated that some materials exhibited a "double hazard”, a large surface area per unit mass as well as a specific surface activity and that "for ranking inflammogenic potency of poorly soluble respirable particles” both properties have to be considered.