J.1.1. Bacterial Mutagenicity Assay

Testing was performed as reported by Zeiger et al.⁷⁵ Glycidamide was sent to the testing laboratory and coded prior to testing. It was incubated with the Salmonella typhimurium tester strains (TA98, TA100) either in buffer or S9 mix (metabolic activation enzymes and cofactors from Aroclor 1254-induced male Sprague Dawley rats) for 20 minutes at 37°C. Top agar supplemented with l-histidine and d-biotin was added, and the contents of the tubes were mixed and poured onto the surfaces of minimal glucose agar plates. Histidine-independent mutant colonies arising on these plates were counted following a 2-day incubation at 37°C.

Each trial consisted of triplicate plates of concurrent positive and negative controls and of at least five doses of glycidamide. In the absence of toxicity, the high dose was limited to 10,000 μg/plate. Trials that gave a positive response were repeated.

In this assay, a positive response is defined as a reproducible, dose-related increase in histidine-independent (revertant) colonies in any one strain/activation combination. An equivocal response is defined as an increase in revertants that is not dose-related, is not reproducible, or is not of sufficient magnitude to support a determination of mutagenicity. A negative response is obtained when no increase in revertant colonies is observed following chemical treatment. There is no minimum percentage or fold-increase required for a chemical to be judged positive or weakly positive.