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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
13. March to 18. April 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: according to EC Directive 92/69/EEC and Regulation EC/440/2008 guideline methods under GLP conditions. This data valid as documented CCRF for this category (Acetylenic geminalic diols).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Lot#: 12671
Purity: 98.3 %

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
mammalian liver post-mitrochondrial fraction (S-9)
Test concentrations with justification for top dose:
see also table in attached report on page -12- regarding "Materials"/Test article".
1.6 to 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: 2-nitrofluorene 5µl, sodium azide 2 µg, 9-aminoacridine 50 µg, 4-nitroquinoline-1-oxide 2µg, Benzo[a]pyrene 10µg or 2-aminoanthracene 5 µg
Details on test system and experimental conditions:
EnviroGem AD01 was assayed for mutation in four histidine-requiring strains of Salmonella typhimurium (TA98, TA100, TA1535, and TA1537), and one tryptophan-requiring strain of Escherichia coli (WP2 uvrA), both in the absence and in the presence of metabolic activation by an Aroclor 1254-induced rat liver post-mitochondrial fraction (S-9), in two separate experiments.

An initial toxicity Range-Finder Experiment was carried out in strain TA100 only, in the absence and presence of S-9, using final concentrations of EnviroGem AD01 at 1.6, 8, 40, 200, 1000, and 5000 mg/plate, plus negative (solvent) and positive controls. Evidence of toxicity was observed following the top dose treatment in the absence and presence of S-9. Strain TA100 treatments were repeated in Experiment 1 in order to investigate the reproducibility of a statistically significant increase in revertant numbers following Range-Finder Experiment treatments in the absence of S-9.

Experiment 1 treatment of all the test strains in the absence and presence of S-9 retained the same test doses as employed for the Range-Finder Experiment. Evidence of toxicity was observed following the top one or two dose treatments of all strains in the absence and presence of S-9, except strain WP2 uvrA in the absence of S-9 where no clear evidence of toxicity was observed.

Experiment 2 treatment of strain TA1535 in the presence of S-9 and strain WP2 uvrA in the absence and presence of S-9 were performed up to 5000 mg/plate. All other treatments in this experiment were performed up to 2000 mg/plate (an estimate of the lower limit of toxicity). In each case narrowed dose intervals were used, in order to more closely investigate those doses of EnviroGem AD01 approaching the limit dose level, and therefore considered most likely to provide evidence of any mutagenic activity. In addition, treatments in the presence of metabolic activation were further modified by the inclusion of a pre-incubation step, in this way the range of mutagenic chemicals that can be detected in this system was increased. Evidence of toxicity was observed following the top dose treatment of all strains in the absence of S-9 (except strain WP2 uvrA where no toxicity was observed), and following the top three dose treatments of all strains in the presence of S-9.

The test article was completely soluble in the aqueous assay system at all concentrations tested, in each of the experiments performed.

Negative (solvent) and positive control treatments were included for all strains in each experiment. The mean numbers of revertant colonies on negative control plates all fell within acceptable ranges, and were significantly elevated by positive control treatments.

No dose-related and reproducible increases in revertant numbers were observed following any strain treatments in the absence or presence of metabolic activation, and therefore this study was considered to have provided no evidence of any EnviroGem AD01 mutagenic activity.

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RANGE-FINDING/SCREENING STUDIES: See details above in "Details in Test System and Conditions"

Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

No dose-related and reproducible increases in revertant numbers were observed following any strain treatments in the absence or presence of metabolic activation, and therefore this study was considered to have provided no evidence of any EnviroGem AD01 mutagenic activity.

It was concluded that EnviroGem AD01 did not induce mutation in four histidine-requiring strains of Salmonella typhimurium (TA98, TA100, TA1535, and TA1537), and one tryptophan-requiring strain of Escherichia coli (WP2 uvrA) when tested under the conditions of this study. These conditions included treatments at concentrations either up to 5000 mg/plate, or the lower limit of toxicity, in the absence and in the presence of a rat liver metabolic activation system (S-9).

It was concluded that EnviroGem AD01 did not induce mutation in four histidine-requiring strains of Salmonella typhimurium (TA98, TA100, TA1535, and TA1537), and one tryptophan-requiring strain of Escherichia coli (WP2 uvrA) when tested under the conditions of this study. These conditions included treatments at concentrations either up to 5000 mg/plate, or the lower limit of toxicity, in the absence and in the presence of a rat liver metabolic activation system (S-9).



Executive summary:

EnviroGem AD01 was assayed for mutation in four histidine-requiring strains of Salmonella typhimurium (TA98, TA100, TA1535, and TA1537), and one tryptophan-requiring strain of Escherichia coli (WP2 uvrA), both in the absence and in the presence of metabolic activation by an Aroclor 1254-induced rat liver post-mitochondrial fraction (S-9), in two separate experiments.

An initial toxicity Range-Finder Experiment was carried out in strain TA100 only, in the absence and presence of S-9, using final concentrations of EnviroGem AD01 at 1.6, 8, 40, 200, 1000, and 5000 mg/plate, plus negative (solvent) and positive controls. Evidence of toxicity was observed following the top dose treatment in the absence and presence of S-9. Strain TA100 treatments were repeated in Experiment 1 in order to investigate the reproducibility of a statistically significant increase in revertant numbers following Range-Finder Experiment treatments in the absence of S-9.

Experiment 1 treatment of all the test strains in the absence and presence of S-9 retained the same test doses as employed for the Range-Finder Experiment. Evidence of toxicity was observed following the top one or two dose treatments of all strains in the absence and presence of S-9, except strain WP2 uvrA in the absence of S-9 where no clear evidence of toxicity was observed.

Experiment 2 treatment of strain TA1535 in the presence of S-9 and strain WP2 uvrA in the absence and presence of S-9 were performed up to 5000 mg/plate. All other treatments in this experiment were performed up to 2000 mg/plate (an estimate of the lower limit of toxicity). In each case narrowed dose intervals were used, in order to more closely investigate those doses of EnviroGem AD01 approaching the limit dose level, and therefore considered most likely to provide evidence of any mutagenic activity. In addition, treatments in the presence of metabolic activation were further modified by the inclusion of a pre-incubation step, in this way the range of mutagenic chemicals that can be detected in this system was increased. Evidence of toxicity was observed following the top dose treatment of all strains in the absence of S-9 (except strain WP2 uvrA where no toxicity was observed), and following the top three dose treatments of all strains in the presence of S-9.

The test article was completely soluble in the aqueous assay system at all concentrations tested, in each of the experiments performed.

Negative (solvent) and positive control treatments were included for all strains in each experiment. The mean numbers of revertant colonies on negative control plates all fell within acceptable ranges, and were significantly elevated by positive control treatments.

No dose-related and reproducible increases in revertant numbers were observed following any strain treatments in the absence or presence of metabolic activation, and therefore this study was considered to have provided no evidence of any EnviroGem AD01 mutagenic activity.

It was concluded that EnviroGem AD01 did not induce mutation in four histidine-requiring strains of Salmonella typhimurium (TA98, TA100, TA1535, and TA1537), and one tryptophan-requiring strain of Escherichia coli (WP2 uvrA) when tested under the conditions of this study. These conditions included treatments at concentrations either up to 5000 mg/plate, or the lower limit of toxicity, in the absence and in the presence of a rat liver metabolic activation system (S-9)