Endo-8-isopropyl-8-azabicyclo[3.2.1]octan-3-ol

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04 April - 03 September 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Endo-8-isopropyl-8-azabicyclo[3.2.1]octan-3-ol

EC Number:

222-314-9

EC Name:

Endo-8-isopropyl-8-azabicyclo[3.2.1]octan-3-ol

Cas Number:

3423-25-4

Molecular formula:

C10H19NO

IUPAC Name:

Endo-8-isopropyl-8-azabicyclo[3.2.1]octan-3-ol

Test material form:

solid: particulate/powder

Method

Target gene:

Salmonella tryphimurium strains TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

S9 Mix

Test concentrations with justification for top dose:

Since no pre-experiment was conducted 5000.0 µg/plate of the test item was selected as the maximum concentration.
The concentration range covered two logarithmic decades.
Two experiments were performed.
According to the dose-selection criteria, the test item was tested at the following concentrations in both experiments independently performed:
31.6; 100.0; 316.2; 1000.0; 2500.0 and 5000.0 µg/plate

Vehicle / solvent:

Positive Control: Without metabolic activation
-Vehicle/solvent used for S. typhimurium TA 100: Aqua dest.
-Vehicle/solvent used for S. typhimurium TA 98: DMSO

Positive Control: With metabolic activation
-Vehicle/solvent used for S. typhimurium TA 98 and TA 100: DMSO

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

other:

Details on test system and experimental conditions:

For the plate incorporation method, the following materials were mixed in a test tube and poured over the surface of a minimal agar plate:
100 µl
- Test solution at each dose level, solvent or negative control or reference mutagen solution (positive control),
500 µl
- S9 mix (for the test with metabolic activation) or S9 mix substitution buffer (for test without metabolic activation),
100 µl
- Bacteria suspension (cf. Preparation of Bacteria, pre-culture of strain),
2000 µl
- Overlay agar.

For the pre-incubation method 100 µl of the test item solution was pre-incubated with the tester strains (100 µl) and sterile buffer or the metabolic activation system (500 µl) for 60 minutes at 37°C for at least 48 h in the dark.

Rationale for test conditions:

The Mutation factor is calculated by dividing the mean value of the revertant counts through the mean values of the solvent control (the exact and not the rounded values are used for calculation).

A test is considered acceptable if for each strain:
- the bacteria demonstrate their typical responses to crystal violet and ampicillin
- the control plates without S9 mix are within the following ranges (range of spontaneous reversion frequencies, historical control data range):
TA 98: 18-63
TA 100: 79-197

- corresponding background growth on both negative control and test plates is observed.
- the positive controls show a distinct enhancement over the control plate.

Evaluation criteria:

A test item is considered as mutagenic if:
- a dose-related increase in the number of relevant occurs and/or
- a reproducible biologically relevant positive response for at least one of the test points occurs in at least one strain with or without metabolic activation.

According to the new OECD guidelines, the biological relevance of the results will be the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.

A test item producing neither a dose related increase in the number of revertants nor a reproducible biologically relevant positive response at any of the test points is consider non-mutagenic in this system.

A biologically relevant increase is described as follows:
- if in test strain TA 100 the number of reversions is at least twice as high
- if in test strain TA 98 the number of reversions is at least three times higher as compared to the spontaneous reversion rate.

Statistics:

The colonies were counted using an Artek-Counter Model 880 (Artek Systems Corporation, BIOSYS GmbH, D-61184 Karben) at maximum sensitivity. If precipitation of the test item precluded automatic counting the revertant colonies were counted by hand.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not
cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used.