Statistical treatment of cytotoxicity in Ames bacterial reverse mutation assays can provide additional structure–activity relationship information

Abstract

The bacterial reverse mutation assay, that is, the Ames test, measures mutations that reverse the inactivation of a gene involved in the synthesis of either histidine in Salmonella bacteria or tryptophan in Escherichia coli. The classic dose–response curve of an Ames assay plots number of reverse mutations (“revertants”) on the y-axis versus dose of the test chemical on the x-axis. Frequently, the dose–response curve resembles a parabola with a linear initial slope resulting from the accumulation of mutations, which transitions to a downward curvature resulting from cell killing (cytotoxicity) at increasingly higher doses of the test chemical. For regulatory purposes, a positive Ames test is usually considered as induction of twice the number of reverse mutations above background levels. For research purposes, the potency of the mutagenic response can be calculated from measuring the initial slope of the mutagenic response. This initial slope can be calculated in a manner that disentangles the downward pull on the initial slope value provided by the initiation of cytotoxicity. For a dose–response curve resembling a parabola, both the initial positive slope representing mutagenicity and the secondary negative slope representing cytotoxicity can be calculated from the same dose–response curve. The Ames test is the most commonly conducted genotoxicity assay. When a series of molecular congeners are assayed in the Ames test for mutagenicity, additional consideration of the cytotoxicity can provide important structure–activity relationship information.