5-methoxy-N-propyl-1,2,3,4-tetrahydronaphthalen-2-aminium chloride

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:

31 May 2012 - 07 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

(2R)-5-methoxy-N-propyl-1,2,3,4-tetrahydronaphthalen-2-amine hydrochloride

Cas Number:

93601-85-5

Molecular formula:

C14H21NO.HCl

IUPAC Name:

(2R)-5-methoxy-N-propyl-1,2,3,4-tetrahydronaphthalen-2-amine hydrochloride

Constituent 2

Reference substance name:

(2S)-5-methoxy-N-propyl-1,2,3,4-tetrahydronaphthalen-2-amine hydrochloride (1:1)

EC Number:

695-797-0

Cas Number:

93601-86-6

Molecular formula:

C14H21NO.HCl

IUPAC Name:

(2S)-5-methoxy-N-propyl-1,2,3,4-tetrahydronaphthalen-2-amine hydrochloride (1:1)

Test material form:

solid: particulate/powder

Details on test material:

- Appearance: Mauve powder
- Storage condition of test material: At room temperature in the dark

Method

Target gene:

- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

Preliminary test (without and with S9) TA100 and WP2uvrA: 3, 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate
Main study: TA1535, TA1537 and TA98:
Without and with S9-mix: 33, 100, 333, 1000, 3330 and 5000 µg/plate

Vehicle / solvent:

- Vehicle used: ethanol
- At concentrations of 10 mg/mL and higher the test substance was suspended in ethanol. At concentrations of 3.33 mg/mL and lower the test substance was dissolved in ethanol. The stock solution was treated with ultrasonic waves to obtain a homogeneous suspension. Test substance concentrations were used within 2 hours after preparation.

Controls

Negative solvent / vehicle controls:

yes

Remarks:

(ethanol)

Positive controls:

yes

Positive control substance:

other: see section "Any other information on materials and methods incl. tables"

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hour

NUMBER OF REPLICATIONS
- Doses of the test substance were tested in triplicate in each strain.

NUMBER OF CELLS EVALUATED: 10E8 per plate

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies.

OTHER EXAMINATIONS:
- The presence of precipitation of the test compound on the plates was determined.

Evaluation criteria:

A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is not greater than two (2) times the concurrent vehicle control, and the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is not greater than three (3) times the concurrent vehicle control.

A test substance is considered positive (mutagenic) in the test if:
a) The total number of revertants in the tester strain TA100 is greater than two (2) times the concurrent vehicle control, or the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is greater than three (3) times the concurrent vehicle control.
b) In case a repeat experiment is performed when a positive response is observed in one of the tester strains, the positive response should be reproducible in at least one independently repeated experiment.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

other: yes/no (see below for details per strain), and tested up to limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

E. coli WP2 uvr A

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

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitation was observed up to and including the top dose of 5000 µg/plate

RANGE-FINDING/SCREENING STUDIES
- In tester strain WP2uvrA, no toxicity or mutagenicity was observed up to and including the top dose of 5000 µg/plate
- In tester strain TA100, toxicity was observed at dose levels of 3330 μg/plate and above in the absence of S9-mix and at 5000 μg/plate in the presence of S9-mix.

COMPARISON WITH HISTORICAL CONTROL DATA
- The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- In tester strain TA98, no toxicity or mutagenicity was observed up to and including the top dose of 5000 µg/plate
- In tester strain TA1535, toxicity was observed at 5000 µg/plate in the absence of S9-mix
- In tester strain TA1537, toxicity was observed at 5000 µg/plate in the absence of S9-mix and at dose levels of 3330 µg/plate and above in the presence of S9-mix

Applicant's summary and conclusion

Conclusions:

The mutagenic activity of the substance was evaluated in accordance with OECD 471 (1997) and according to GLP principles. Based on the results of this study it is concluded that the substance is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay with and without metabolic activation.

Executive summary:

The mutagenic activity of the substance was evaluated in accordance with OECD 471 (1997) and according to GLP principles. The substance was tested up to and including 5000 µg/plate in all tester strains, in the absence and presence of S9-mix. Adequate solvent and positive controls were included. The substance did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four S. typhimurium tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA, both in the absence and presence of S9-metabolic activation. Based on the results of this study it is concluded that the substance is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay with and without metabolic activation.