Tests forward mutation

The mouse lymphoma forward mutation test at the thymidine kinase locus (detects mutations to a nonfunctional thymidine kinase in a line of culture mouse lymphoma cells). 

Forward Mutation Tests. Forward mutation is an endpoint that may arise from various events, including base substitutions, frameshifts, DNA deletions, and so on, as mentioned earlier. 

Han, J.-S. (1997) Mutagenic activity and specificity of hydrogen peroxide in the ad-3 forward-mutation test in two-component heterokaryons of Neurospora crassa. Mutat. Res., 374, 169-184... 

In the forward mutation test, bacteria are added to a medium containing a harmful substance (eg. 8-azaguanine). Under these conditions most of the bacteria will die, but some will survive because they have adapted their genotype by mutation. In the forward mutation test the development of resistence (by mutation) above the control mutation frequency is studied in the presence of the test compound. This is also done in the presence and absence of the metabolic activation system. 

In 1985 it was published [1] that stevioside was completely safe but that metabolically activated steviol was mutagenic in a "Forward Mutation Test". Steviol had to be applied together with the microsomal fraction of liver of animals treated with carcinogenic compounds (polychlorinated biphenyl or phenobarbital plus 5,6-benzoflavone). This publication has led to a confusing discussion between advocates and opponents of the use of Stevia or stevioside. To unravel the problem we must consider the opinion of authoritative international organisations such as OECD (Organisation for Economic Co-Operation and Development) and ICH (International Council of Harmonisation). To accept new substances as food three different mutagenesis tests are accepted and required by the OECD as well as by the ICH. These can be seen in Table 1. 

In the Ames test, mouse lymphoma cell (L5178Y/ TK ") forward mutation test, Chinese Hamster Ovary (CHO) cell chromosome aberration test, and mouse micronucleus test, alvimopan was not found genoto-xic. Its metabolite was negative in the Ames test, CHO cell chromosome aberration test, and mouse micronucleus test. 

Macgregor, J.T., D.H. Gould, A.D. Mitchell, and G.P. Sterling. 1979. Mutagenicity tests of diflubenzuron in the micronucleus test in mice, the L5178Y mouse forward mutation assay, and the Ames Salmonella reverse mutation test. Mutat. Res. 66 45-53. 

The test is used to detect forward and reverse gene mutation in Aspergillus nidulans, a eukaryotic fungus. These mutations are detected by changes in colonial morphology or nutritional requirements in treated populations. The methionine and 2-thioxanthine forward mutation systems can be used to detect mutations in Aspergillus nidulans. 

A variety of mammalian cell culture systems can be used to detect mutations induced by chemical substances. The L5178Y mouse l)nnphoma line, measuring mutation at the TK locus, is preferred. TK is an important enz)une involved in DNA synthesis. Cells are exposed to the test substance at various concentrations, in the presence and absence of a metabolic activation system, for a suitable period of time, and then subcultured to assess cytotoxicity and to allow phenotypic expression prior to mutant selection. Cells deficient in TK because of a forward mutation are resistant to the cytotoxic effects of pyrimidine analogues (antimetabolites), such as trifluorothymidine (TFT). This is because the antimetabolites cannot be incorporated into cellular nucleotides and kill the cell through inhibition of cellular metabolism. After treatment, cells are grown in a medium containing TFT mutant cells can proliferate in the presence of TFT, whereas normal cells containing TK are killed. This allows the detection of an increase in mutant... 

The genotoxic potential of brinzolamide was evaluated by the bacterial reverse mutation assay (Ames test) with S typhimurium and E coli, an in vitro mouse lymphoma forward mutation assay (MLFMA), an in vivo... 

As with the Salmonella reversion assay, this shortterm test is conducted both without (— PMS) and with metabolic activation produced by addition of post-mitochondrial supernatant containing rat liver enzymes ( + PMS). These terms are equivalent to — S9 and + S9 in the Ames reversion assay we use the latter designation for both types of bacterial assays. A more sensitive micro-forward mutation bioassay using this TM677 strain to determine the mutagenicity of indoor air particles, including ETS and wood smoke, is described by Lewtas et al. (1987). 

The human cell mutagenicities of several mono- and dinitropyrenes were determined by Busby and coworkers (1994b) in a forward mutation assay using a metabolically competent line of MCL-5 cells. Minimum detectable mutagen concentrations (MMC, in nmol ml-1 the smaller the detectable concentration, the greater the potency) were as follows 1,6-dinitropyrene, 0.8 1,8-dinitropyrene, 1.5 4-nitropyrene, 3.1 1-nitro-pyrene, 9.1. Other PACs tested, 2-nitropyrene, 1,3-dinitropyrene, and pyrene, were nonmutagens in this... 

Oberly, T.J., Bewsey, B.J. Probst, G.S. (1985) Tests for the induction of forward mutation at the thymidine kinase locus of L5178Y mouse lymphoma cells in culture. Prog. Mutat. Res., 5, 569-582... 

Careie, A., Conti, G, Conti, L. Crebelli, R. (1985) Assays in Aspergillus nidulans for the indnction of forward-mutation in haploid strain 35 and for mitotic nondisjunction, haploidization and crossing-over in diploid strain PI. In Ashby, J., de Serres, F.J., Draper, M., Ishidate, M., Jr, Margolin, B.H., Matter, B.E. Shelby, M.D., eds. Progress in Mutation Research, Volume 5, Evaluation of Short-Term Tests for Carcinogens. Report of the International Programme on Chemical Safety s Collaborative Study on in vitro assays, Amsterdam, Elsevier Science, pp. 307-312... 

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