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Effect of storage at 4 degrees on prepared Salmonella/microsome test plates
Abstract:
Salmonella/microsome soft-agar overlay ('Ames' test) plates were prepared using the previously published 'York' method. Plates treated with either added 3-methylcholanthrene (3MC) or 3,3'-dichlorobenzidine (DCB) and Aroclor-induced rat-liver S9 were stored at 4 degrees after preparation and removed at time intervals thereafter for incubation at 37 degrees. The number of 3MC-induced mutants in TA100 fell within 24 h of storage by 50% but then remained at this level until 96 h. Storage for a total of 336 h still showed some residual mono-oxygenase activity. On the other hand the ability to convert DCB to a mutagen for TA98 appeared to increase with storage, reaching a peak by 96 h. After this time the number of induced mutants fell until by 336 h the numbers were approximately equal to those plates which had not been stored in the cold.
Insights
Storage affects the mutagenicity of compounds in the Ames test. 3-methylcholanthrene mutagenicity decreased over time, while 3,3'-dichlorobenzidine mutagenicity initially increased before decreasing.
Area of Science:
- Toxicology
- Genetics
- Biochemistry
Background:
- The Ames test is a standard method for assessing chemical mutagenicity.
- Understanding the stability of metabolic activation systems is crucial for accurate mutagenicity testing.
Purpose of the Study:
- To investigate the impact of cold storage duration on the mutagenic potential of chemicals using the Salmonella/microsome assay.
- To evaluate the stability and activity of rat-liver S9 enzyme preparations under storage conditions.
Main Methods:
- Salmonella/microsome (Ames) test plates were prepared using the York method.
- Plates were treated with 3-methylcholanthrene (3MC) or 3,3 -dichlorobenzidine (DCB) and Aroclor-induced rat-liver S9.
- Plates were stored at 4°C for varying durations (up to 336 hours) before incubation at 37°C.
Main Results:
- Mutagenicity induced by 3MC in strain TA100 decreased by 50% within 24 hours of storage and remained stable up to 96 hours.
- Mono-oxygenase activity showed residual presence even after 336 hours of storage.
- The mutagenic activation of DCB for strain TA98 increased with storage, peaking at 96 hours, then declining by 336 hours.
Conclusions:
- Cold storage duration significantly alters the mutagenic response in the Ames test, depending on the specific chemical and metabolic activation pathway.
- The stability of S9 enzyme activity varies, impacting the accurate assessment of mutagenicity for different compounds over time.