Abstract
The fluctuation test is a means of measuring bacterial mutation in a liquid medium. It has been modified such that it may be used in several different ways using different types of metabolising systems. The incorporation of isolated hepatocytes instead of S9 fraction for metabolic activation has been developed to provide suitable culturing conditions for the hepatocytes and bacteria. The test has been used to look at the mutagenicity of 2AAF with S9 derived from differently pretreated animals and also isolated hepatocytes. It was observed that there is an S9 optimum for maximal mutagenic activity of 2AAF, which is very much lower with aroclor-induced S9 than uninduced S9 in both the fluctuation test and the Ames test. The optimal S9 concentration is greater for both aroclor-induced and uninduced S9 in the Ames test than in the fluctuation test indicating sane differences between these two tests. As a screening test, the fluctuation test has been validated in an International study using carcinogen and structurally related non-carcinogen pairs. Using uninduced S9 and uninduced hepatocytes the test has a high predictivity of carcinogenicity. However, hepatocytes alone offer little advantage over S9 fraction for general screening purposes. When using hepatocytes in comparison with S9, NM, DMBA and DMA showed increased mutagenicity, while unexpectedly 2AAF showed reduced mutagenicity. 2NA and 1NA were not mutagenic with hepatocytes. In general, where a mutagenic response was observed with S9 fraction there was a mutagenic response with hepatocytes. Hepatocytes are likely to be more useful as a research tool than in screening tests. The fluctuation test has also been used to assess the mutagenicity of some synthetic phenol metabolites and their sulphate esters of BP. 1-OHBP, 3-OHBP and 12-OHBP were mutagenic without metabolic activation, while 9-OHBP and 7-OHBP were mutagenic with aroclor-induced S9, but not with 3MC-induced Ca[2+] precipitated microsomes. The sulphate esters of 3-, 7- and 9-OHBP were clearly mutagenic in the presence of aroclor-induced S9. The mutagenicity of BP was increased by the presence of the synthetic steroid betamethasone using aroclor-induced S9, while very little effect was observed with uninduced S9. The use of the fluctuation test to assess the mutagenicity of complex mixtures of chemicals was exemplified by a study on several oils.