Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Chromosome abberation Bis(4-(1,1,3,3-tetramethylbutyl) phenyl) amine is negative in human lymphocytes and is considered to be non-clastogenic and non-aneugenic to human lymphocytes (Harlan Laboratories, 2013). Mammilian gene mutation Bis(4-(1,1,3,3-tetramethylbutyl) phenyl) amine is negative in mouse lymphoma L5178Y cells and is not considered mutagenic in the mouse lymphoma L5178Y test system (Harlan Laboratories., 2013). Ames Bis(4-(1,1,3,3-tetramethylbutyl) phenyl) amine is negative in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 (with and without activation) and is considered to be non-mutagenic to bacteria (E. Coli WP2 uvrA) (Harlan Laboratories Ltd, 2012).

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 21 September 2012 to 22 January 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted in accordance with validated testing guidelines and under GLP conditions.

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian cell gene mutation assay

Species / strain / cell type:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

Experiment 1: 15.63, 31.25, 62.5, 125, 250, 500 μg/mL
Experiment 2: 15.63, 31.25, 62.5, 125, 250, 500 μg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone

No additional data

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

Experiment 1 (-S9) 4-Hour Exposure; Experiment 2 (-S9) 24-Hour Exposure

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

Experiment 1 (+S9) 4-Hour Exposure

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: In Experiment 1, 4-hour exposure; In Experiment 2, 4-hour exposure and 24-hour exposure

NUMBER OF REPLICATIONS: 2

OTHER EXAMINATIONS:
- Other: The daily cell counts were used to obtain a Relative Suspension Growth (%RSG) value that gives an indication of post treatment toxicity during the expression period as a comparison to the vehicle control, and when combined with the Viability (%V) data a Relative Total Growth (RTG) value.

No additional data

Evaluation criteria:

Vehicle controls results should ideally be within this range, although minor errors in cell counting and dilution or exposure to the metabolic activation system may cause this to be slightly elevated. Experiments where the vehicle control values are markedly greater than 2E-04 mutant frequency per survivor are
not normally acceptable and will be repeated.

Positive control chemicals should induce at least three to five fold increases in mutant frequency greater than the corresponding vehicle control.

Statistics:

None stated

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not valid

Positive controls validity:

valid

Additional information on results:

With no evidence of significant toxicity in the preliminary toxicity test, the maximum dose level used in the Mutagenicity Test was limited by the onset of greasy/oily precipitate that was considered to be effectively reducing exposure of Bis(4‐(1,1,3,3‐tetramethylbutyl)phenyl)amine to the cells.

In the Mutagenicity Test, precipitate of Bis(4‐(1,1,3,3‐tetramethylbutyl)phenyl)amine was observed at and above 31.25 μg/mL and became greasy/oily in appearance at 500 μg/mL. The vehicle (solvent) controls had mutant frequency values that were considered acceptable for the L5178Y cell line at the TK +/- locus.

The positive control items induced marked increases in the mutant frequency indicating the satisfactory performance of the test and of the activity of the metabolising system.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Conclusions:

Interpretation of results (migrated information):
negative

Bis(4‐(1,1,3,3‐tetramethylbutyl)phenyl)amine did not induce any toxicologically significant increases in the mutant frequency at the TK +/- locus in L5178Y cells and is therefore considered to be non-mutagenic under the conditions of the test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

Ames

Key study:

According to an OECD 471 conducted according to GLP, Bis(4-(1,1,3,3-tetramewthylbutyl)phenyl)amine is negative in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 (with and without activation). Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine is considered to be non-mutagenic to bacteria under the conditions of this test (Harlan Laboratories Ltd, 2012).

Chromosome aberration

Key Study:

According to an OECD 476, conducted according to GLP, Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine is negative in human lymphocytes. Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine is considered to be non-clastogenic and non-aneugenic to human lymphocytesin vitro (Harlan Laboratories Ltd, 2013).

Mammilian gene mutation

According to an OECD 476, conducted according to GLP, Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine is negative using mouse lymphoma L5178Y cells. Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions (Harlan Laboratories Ltd., 2013).

Justification for selection of genetic toxicity endpoint
The study was conducted according to appropriate OECD Guidelines (OECD 487), under GLP conditions at a reputable European Union laboratory.
The study in mammalian cells was chosen over that in bacterial cells, as the higher level study.

Justification for classification or non-classification

Bis(4 -(1,1,3,3 -tetramethylbutyl)phenyl)amine is negative in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 (with and without activation), negative in human lymphocytes and negative using mouse lymphoma L5178Y cells.

Based upon Table 3.5.1 of Regulation EC No. 1272/2008 on classification, labelling and packaging (CLP) of substances and mixtures, Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine is not classified for genotoxicity.