Acute inhalation toxicity of volatile organic compounds - application of an improved cell-based in vitro procedure

Cell based in vitro methods to assess the biological effects of inhalable compounds are based on "air-lifted interface" (ALI) cell culture protocols where cultures from cell lines, primary cells or ex vivo sources such as PCLS (precision cut lung slices) are exposed efficiently to airborne material. They have been applied in approaches to test toxicity and biological action of environmental, workplace or chemical compounds. The basic ALI procedure has been successfully prevalidated as a short-exposure test (1 hour exposures) for chemical gases. However, although a good predictability for highly toxic gaseous compounds or non-toxic inert gases could be demonstrated, it was not clear until now, if highly hydrophobic or low-toxic volatile organic compounds (VOCs) can also be assessed by cell based methods in vitro under these conditions with regard to a relevant estimate of their acute inhalation toxicity characteristics in vivo. Therefore, a four compound test substance matrix was set up from the data base of the European chemicals agency (ECHA) including VOCs differing in hydrophobicity and acute in vivo inhalation toxicity. A cell based testing including vaporized compounds, an improved air-liquid interface in vitro procedure (PRIT ExpoCube) with human lung cells (A549) and read-out of cytotoxicity (WST-1) was carried out. In vitro toxicity data were compared to acute in vivo inhalation toxicity data from the ECHA data base. In vitro toxicity data from 1,1-dimethylethyl hydroperoxide (hydrophilic, high acute in vivo inhalation toxicity), 2-methylpropan-1-ol (hydrophilic, low toxicity), toluene (hydrophobic, low toxicity) and styrene (hydrophobic, low toxicity) could be derived from all substances and reflected the relative high and low acute in vivo inhalation toxicity potency. The results demonstrate that it was possible to study the acute inhalation toxicity by cell based methods even for low toxic, hydrophobic airborne chemicals nd thereby characterize the in vitro approach as a useful alternative method for screening purposes in the sense of the "3Rs".