Decamethylenediamine

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2008-07-29 to 2008-08-08

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study; GLP study without deviations

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

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 (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Decamethylenediamine

EC Number:

211-471-9

EC Name:

Decamethylenediamine

Cas Number:

646-25-3

Molecular formula:

C10H24N2

IUPAC Name:

decane-1,10-diamine

Constituent 2

Reference substance name:

1, 10-Decanediamine

IUPAC Name:

1, 10-Decanediamine

Test material form:

other: crystalline solid

Details on test material:

1,10-Decanediamine
purity > 99%

Method

Target gene:

mutated gene loci responsible for histidine auxotrophy

Species / strain

Species / strain 1

Species / strain / cell type:

E. coli WP2

Additional strain / cell type characteristics:

other: Tryptophan-independent

Species / strain 2

Species / strain / cell type:

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

Additional strain / cell type characteristics:

other: histidine auxotroph

Metabolic activation:

with and without

Metabolic activation system:

Phenobarbital/ß-Naphthoflavone induced rat liver S9; male Wistar rats

Test concentrations with justification for top dose:

Pre-Experiment/Experiment I and II:
3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

Vehicle / solvent:

Ethanol

Controls

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

Ethanol

True negative controls:

no

Positive controls:

yes

Remarks:

for details see below

Positive control substance:

other: sodium azide; 4-nitro-o-phenylene-diamine; methyl methane sulfonate; 2-aminoanthracene

Details on test system and experimental conditions:

Bacterial Reverse Mutation Test
SYSTEM OF TESTING
- Pre-Experiment / Experiment I: plate incorporation test (+/- metabolic activation)
Experiment II: pre-incubation assay (+/- metabolic activation)
- Metabolic activation assay: Phenobarbital/ß-Naphthoflavone induced Wistar rat liver S9 (protein concentration in S9: 30.7 mg/ml)
ADMINISTRATION
- Dosing: Pre-Experiment and Experiment I/II : 3 - 5000 µg/plate
- Number of replicates: 3
- Positive and negative control groups and treatment:
- without metabolic activation:
sodium azide for TA 1535 and TA 100
4-nitro-o-phenylene-diamine for TA 1537 and TA 98
methyl methane sulfonate for WP2 uvrA
- with metabolic activation:
2-aminoanthracene for all strains
- negative control: untreated; solvent control: Ethanol
- Pre-incubation time: 60 min at 37 °C; incubation time: 48 h at 37 °C in the dark

Evaluation criteria:

CRITERIA FOR EVALUATING RESULTS:
mutagenic effects (i.e ratio of revertant rates treated/control >=2) at <= 5000µg/plate with generally positive dose-response relationship in any
strain

Statistics:

According to the OECD Guideline 471, a statistical analysis of the data is not mandatory

Results and discussion

Test results

Species / strain:

other: S. typhimurium TA 98; TA 100; TA1535 and TA 1537; E. coli WP2 uvrA

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

see below

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

GENTOXIC EFFECTS:
- With metabolic activation: None (even at cyctotoxic concentration)
- Without metabolic activation: None (even at cytotoxic concentration)
PRECIPITATION CONCENTRATION: > 5000 µg/plate
CYTOTOXIC CONCENTRATION: see remarks on results

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

The plates incubated with the test item showed reduced background growth at the following concentrations (µg/plate):

Strain Experiment I Experiment II

without S9 mix       with S9 mix without S9 mix       with S9 mix

TA 1535 2500 -5000       2500 -5000 2500 -5000              5000

TA 1537        2500 -5000 2500 -5000 2500 -5000 5000

TA 98 1000 -5000 2500 -5000                   2500 -5000 2500 -5000

TA 100               1000 -5000        2500 -5000               2500 -5000               5000

WP2 uvrA        2500 -5000 2500 -5000                            /                            /

/ = no reduced background growth observed

Toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in the test groups at the following concentrations (µg/plate):

Strain Experiment I                                           Experiment II

without S9 mix with S9 mix              without S9 mix              with S9 mix

TA 1535                     5000              2500 -5000                     /                                       /

TA 1537                     5000                      /                             5000                                   /

TA 98                     1000 -5000       2500 -5000              2500 -5000                         5000

TA 100                   1000 -5000       2500 -5000               2500 -5000                      5000

WP2 uvrA               2500 -5000       2500 -5000                      /                                         /

/ = No toxic effects observed

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
induce gene mutations by base pair changes or frameshifts in the genome of the strains used.
Therefore, decamthylenediamine is considered to be non-mutagenic in this Salmonella typhimurium and Escherichia coli reverse mutation assay.

Executive summary:

This study was performed to investigate the potential of decamethylenediamine to induce gene mutations in the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100, and the Escherichia coli strain WP2 uvrA.

The assay was performed in two independent experiments both with and without liver microsomal activation. No data were evaluated for strains TA 1535 without S9 mix and TA 1537 with and without metabolic activation in experiment I, since the data of the controls did not met the acceptability criteria of the assay. These parts of experiment I were repeated under identical conditions and the results are reported as part of experiment I. Each concentration, including the controls, was tested in triplicate. The test item was tested at the following concentrations: 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

The plates incubated with the test item showed reduced background growth at higher concentrations in all strains.

Toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in the test groups with and without metabolic activation.

No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with 1, 10-Decanediamine at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance.

Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.

In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used. Therefore, decamethylenediamine is considered to be non-mutagenic in this Salmonella typhimurium and Escherichia coli reverse mutation assay.