Evaluation of modified bacterial mutagenicity assays for the genotoxicity testing of mineral oils

Abstract

A modified bacterial mutagenicity assay based on the Ames Salmonella/mammalian microsome test has been developed for application in the genotoxicity testing of mineral oils. The assay uses washed microsomes from rat liver in place of S9 fraction in order to increase the sensitivity of detection of genotoxicity. The modified assay was used to test a series of oils for which skin carcinogenicity bioassay data in mice were available. Oils were tested as emulsions in water using Tween 80 as a dispersant. A mutagenicity index for each oil was obtained using non-linear regression analysis of data from the dose-response curve. The results showed an empirical correlation between increasing mutagenicity index, carcinogenicity and the polycyclic aromatic hydrocarbon content of the oils. The washed-microsome assay was also compared with modified Ames assays developed by Blackburn et al. (Cell Biol. Toxicol., 1, 40, 1984; Cell Biol. Toxicol., 2, 63, 1986) which employed increased levels of S9 (rat and hamster liver) to test dimethyl sulphoxide extracts of oils. The washed-microsome assay can be used for the testing of whole oils rather than extracts which are necessary for the modified Ames assay. It is recognised that the determinants of carcinogenic activity in vivo include promoting activity which such assays are unable to detect. Nevertheless, such modified bacterial assays may be a useful prescreen since genotoxicity is recognised as a key initial step in carcinogenesis.

Scientific Opinion on Mineral Oil Hydrocarbons in Food

“…Most studies on the genotoxicity of mineral oil and mineral oil fractions have been performed in the Salmonella typhimurium mutagenicity assay (Blackburn et al, 1984(Blackburn et al, , 1986IARC, 1984IARC, /1987Roy et al, 1988;Granella and Clonfero, 1991;Brooks et al, 1995;Mackerer et al, 2003). With the exception of highly purified oil varieties consisting of alkanes and naphthenes, all mineral oils are mutagenic in the Salmonella typhimurium assay ( IARC, 1984;Roy et al, 1988;Mackerer et al, 2003).…”

“…Generally, upon testing in the Salmonella typhimurium assay, in spite of their carcinogenic activity, whole mineral oils often show weak genotoxic activity (Brooks et al, 1995). Several approaches have therefore been used to enhance the sensitivity of the bacterial tests.…”

“…These modified methods include prior fractionation by solvent extraction (with DMSO alone or in combination with cyclohexane) in order to concentrate the total polycyclic aromatic hydrocarbon fraction. Also chromatographic separation has been employed for testing of individual components (Epler et al, 1979;Brooks et al, 1995;Mackerer et al, 2003). Since the mineral hydrocarbons are not soluble in water, procedures involving emulsification of oils have also been used (Brooks et al, 1995).…”

“…Also chromatographic separation has been employed for testing of individual components (Epler et al, 1979;Brooks et al, 1995;Mackerer et al, 2003). Since the mineral hydrocarbons are not soluble in water, procedures involving emulsification of oils have also been used (Brooks et al, 1995). A modified Salmonella typhimurium assay for testing was developed by Blackburn et al (1984Blackburn et al ( , 1986 using DMSO extracts of mineral oils in a test system with increased concentration of rat or hamster liver S-9 activating system as well as co-factors.…”

“…BaP may be inhibited by less active MOAH) (Brooks et al, 1995;Booth et al, 1998;Mackerer et al, 2003). Hence, it is not possible to sum up the activity of a number of single fractions.…”