Amaranth suppresses the mutagenicity of 2-acetylaminofluorene by lowering the concentration of nadph in top agar

Addition of 1 mg amaranth (FD&C Red No. 2) to the top agar of Salmonella/S9 assay plates decreased the yield of revertants induced by 20 micrograms 2-acetylaminofluorene (AAF) by over 50% and additional amaranth completely eliminated the mutagenic response. Similar suppression of AAF mutagenicity was seen with sulfonazo III, another azo dye. The suppressive effect of amaranth was greatest at low S9 concentrations and decreased as the amount of S9 was increased. When N-hydroxyacetylaminofluorene (N-OH-AAF) was used as mutagen, amaranth had little or no effect on either the number of revertants obtained or the S9 optimum. Similarly, 1-naphthylamine-4-sulfonic acid (a reduction product of amaranth) did not significantly affect the mutagenicity of AAF. The rate of metabolism of [14C]AAF by the S9 preparations was shown to be markedly decreased by amaranth, as were the levels of both the phenolic metabolites and of N-OH-AAF. Thus, it appeared that amaranth acts by blocking the conversion of AAF to N-OH-AAF and that this effect is caused by the amaranth itself and not by its constituent amines. Further experiments indicated that amaranth greatly decreased the levels of NADPH formed in reaction mixtures comparable to S0 mix in top agar and that such reaction mixtures also metabolized amaranth to colourless compounds. It appears likely that in top agar, NADPH reacts with amaranth at a fast enough rate to limit severely the level of the reduced co-factor (which must be formed from NADPH+ by the action of endogenous glucose-6-phosphate dehydrogenase) and thus decreases the rate of activation of mutagens by other NADPH-dependent processes.