Toxicity characterization of complex mixtures using biological and chemical analysis in preparation for assessment of mixture similarity.

In the United States, several proposed approaches for using bioassays for the risk assessment of complex hazardous mixtures require that selected mixtures be "sufficiently similar" to each other. The goal of this research was to evaluate the utility of a protocol using in vitro bioassays and chemical analysis as a basis for assessing mixture similarity. Two wood preserving wastes (WPWs) containing polycyclic aromatic hydrocarbons and pentachlorophenol were extracted and fractionated to generate potentially similar mixtures. Chemical analysis was conducted using gas chromatography/mass spectrometry. Genotoxicity was evaluated using the Salmonella/microsome and Escherichia coli prophage induction assays. The crude extract of one WPW was also tested in the chick embryotoxicity screening test (CHEST) assay. The CHEST assay provided the most sensitive measurement of toxicity. Overall, the biological potency of the samples was not well correlated with predicted potency based on chemical analysis. Although several mixtures appeared similar based on chemical analysis, the magnitude of the response in bioassays was often dissimilar. Fractionation was required to detect the genotoxicity of mixture components in vitro. The results confirm the need for an integrated protocol, combining chemical analysis, fractionation, and biological testing to characterize the risk associated with complex mixtures.

Toxicity assessment of complex mixtures remains a goal.

One of the initial steps in remediating contaminated environments is to assess the human and ecological health risk associated with exposure to contaminants in a specific medium. Presented here are the results of a five-year study investigating the toxicity of simple and complex mixtures. A series of model compounds and simple mixtures including polycyclic aromatic hydrocarbons (PAHs), pentachlorophenol (PCP), and halogenated aliphatic hydrocarbons (HAHs) were analyzed. Mixture toxicity was studied using microbial genotoxicity assays and cytotoxicity assays with renal and neural cells. The majority of binary mixtures described here induced additive responses. A limited number of samples were identified where binary mixtures induced inhibitory effects. For example, benzo(a)pyrene (BAP) alone induced 30% renal cell death, whereas an equimolar dose of chrysene and BAP only produced 1.6% cellular death. In none of the mixtures tested did the mixture toxicity results deviate from the predicted results by an order of magnitude. The results from testing binary mixtures in this study indicate that the results did not deviate significantly from additivity. Complex mixture results were more difficult to interpret. The toxicity of complex mixtures could not be accurately predicted based on chemical analysis. This could be due to chemical interactions or due to the presence of unidentified chemicals, such as alkyl PAHs or high molecular weight PAHs that are not included in the standard risk assessment procedure. Even though the results from these in vitro studies indicate that additive assumptions will generally be appropriate for binary mixtures similar to the ones tested here, the risk associated with complex mixtures remains a challenge to predict. Before the results of toxicity testing can be used to adjust risk assessment calculations, it is important to fully appreciate the chemical composition and to understand the mechanism of observed chemical interactions in animals chronically exposed to low doses of chemical mixtures. This research was supported by ATSDR Grant no. ATU684505 and NIEHS SBRP Grant no. P42 ES04917.