1-[4-(4-chlorophenoxy)-2-(trifluoromethyl)phenyl]ethanone

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test substance did not show any mutagenic response in the bacterial reverse mutation assay (BASF SE, 40M0500/14M238, 2015).

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-01-13 to 2015-01-23

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

21 July 1997

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

30 May 2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Version / remarks:

August 1998

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

BASF SE, Experimental Toxicology and Ecology, 67056 Ludwigshafen, Germany

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- batch No.of test material: L85-76

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Storage stability: The stability of the test substance under storage conditions was guaranteed until 01 Oct 2016 as indicated by the sponsor, and the sponsor holds this responsibility.

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:

S9 mix (phenobarbital and β-naphthoflavone induced rat liver)

Test concentrations with justification for top dose:

Standard plate test (SPT): 33, 100, 333, 1000, 2600, 5200 µg/plate
Preincubation test (PIT): 33, 100, 333, 1000, 2600, 5200 µg/plate

Vehicle / solvent:

- Vehicle used: DMSO
- Justification for choice of solvent/vehicle: Due to the limited solubility of the test substance in water, DMSO was used as vehicle, which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data are available.

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2- aminoanthracene

Remarks:

with S9 mix (all tester strains tested)

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: N-methyl-N'-nitro-N-nitrosoguanidine

Remarks:

without S9 mix; 5 µg/plate (TA 1535, TA 100)

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylenediamine

Remarks:

Withiut S9 mix; 10 µg/plate (TA 98)

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

Without S9 mix; 100 µg/plate (TA 1537)

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

Without S9 mix; 5 µg/plate (E. coli WP2 uvrA)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation) (SPT); preincubation (PIT)

DURATION
- Preincubation period: 20 min
- Exposure duration: 48 - 72 hours in the dark at 37 °C

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: decrease in the number of his+ or trp+ revertants

Evaluation criteria:

Acceptance criteria
Generally, the experiment was considered valid if the following criteria were met:
• The number of revertant colonies in the negative controls was within the range of the historical negative control data for each tester strain.
• The sterility controls revealed no indication of bacterial contamination.
• The positive control substances both with and without S9 mix induced a distinct increase in the number of revertant colonies within the range of the historical positive control data or above.
• Fresh bacterial culture containing approximately 109 cells per mL were used.

Assessment criteria
The test substance was considered positive in this assay if the following criteria were met:
• A dose-related and reproducible increase in the number of revertant colonies, i.e. at least doubling (bacteria strains with high spontaneous mutation rate, like TA 98, TA 100 and E.coli WP2 uvrA) or tripling (bacteria strains with low spontaneous mutation rate, like TA 1535 and TA 1537) of the spontaneous mutation rate in at least one tester strain either without S9 mix or after adding a metabolizing system.
A test substance was generally considered non-mutagenic in this test if:
• The number of revertants for all tester strains were within the historical negative control data range under all experimental conditions in at least two experiments carried out independently of each other.

Key result

Species / strain:

other: TA 1535, TA 100, TA 1537, TA 98, E. coli WP2 uvrA

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

SPT: Weak bacteriotoxic effect at 2600 µg/plate in tester strain TA 100 with S9 mix and in tester strain E. coli WP2 uvrA without S9 mix. PIT: Bacteriotoxicity at 5200 µg/plate in tester strain TA 1537 with S9 mix.

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No
- Effects of osmolality: No
- Evaporation from medium: No
- Precipitation: Precipitation of the test substance was found at 5200 µg/plate with and without S9 mix.

HISTORICAL CONTROL DATA
- Positive historical control data: Yes (Oct 2013 - Jul 2014)
- Negative (solvent/vehicle) historical control data: Yes (Oct 2013 - Jul 2014)

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- A weak bacteriotoxic effect was occasionally observed depending on the strain und test conditions from about 2600 µg/plate onward.

Experiment 1 (SPT)
Dose [µg/plate]	Mean number of revertant colonies/ 3 replicate plates with different strains of Salmonella typhimurium and Escherichia coli
	TA 1535	TA 100	TA 1537	TA 98	E. coli
	Results without S9 mix
DMSO	10.7	26.0	7.7	23.7	69.3
33	17.0	25.7	8.7	25.7	70.0
100	11.3	33.3	7.7	15.7	54.7
333	13.7	21.7	7.0	30.3	58.3
1000	18.7	31.3	5.7	23.7	49.3
2600	14.7	25.3	9.0	24.3	23.0
5200	16.3	38.7	8.7	17.3	55.7
Positive control	4879.3	4306.3	496.0	369.3	927.3
	With S9 mix
DMSO	15.3	34.0	10.3	25.7	79.0
33	21.3	34.0	10.7	32.2	73.0
100	18.7	43.7	11.0	30.7	71.3
333	17.7	27.0	11.0	26.7	71.0
1000	10.3	23.3	14.0	24.7	56.7
2600	16.3	13.7	11.3	27.3	76.0
5200	16.7	22.7	11.3	27.0	61.7
Positive control	228.3	695.3	175.7	1878.0	254.0

Experiment 2 (PIT)
Dose [µg/plate]	Mean number of revertant colonies/ 3 replicate plates with different strains of Salmonella typhimurium and Escherichia coli
	TA 1535	TA 100	TA 1537	TA98	E. coli
	Without S9 mix
DMSO	8.0	27.7	8.3	25.7	65.7
33	9.7	23.7	7.0	21.7	57.0
100	9.3	29.7	9.3	24.0	60.7
333	9.3	28.3	8.0	22.0	61.3
1000	8.3	30.7	8.3	25.7	52.7
2600	9.0	25.7	9.0	26.0	59.0
5200	11.7	30.3	6.3	26.0	54.0
Positive control	2588.7	3006.7	419.7	385.7	412.3
	With S9 mix
DMSO	9.7	42.0	8.3	30.0	58.3
33	10.0	37.0	9.3	28.0	70.3
100	8.3	39.3	10.7	27.7	66.7
333	9.0	41.3	6.7	27.0	58.7
1000	11.7	41.0	7.0	27.7	66.7
2600	9.7	44.3	8.3	34.0	63.3
5200	9.7	41.7	2.3	29.3	66.0
Positive control	170.7	534.7	80.3	961.7	148.0

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

In a reverse mutation assay in bacteria according to OECD guideline 471, strains of S. typhimurium (TA1535, TA1537, TA98, TA100) and E. coli (WP2 uvr A) were exposed to the test substance at concentrations of 33, 100, 333, 1000, 2600, 5200 µg/plate) in the presence and absence of metabolic activation (S9 mix) (BASF SE,40M0500/14M23, 2015). A standard plate incorporation test and a pre-inucbation assay were performed. Three test plates per dose were tested.
The test substance was tested up to the limit concentration of 5200 µg/plate. Precipitation of the test substance was found at 5200 µg/plate with and without S9 mix. A weak bacteriotoxic effect was occasionally observed depending on the strain and test conditions from about 2600 µg/plate onward.
According to the results of the present study, the test substance did not lead to an increase in the number of revertant colonies either without S9 mix or after adding a metabolizing system in two experiments carried out independently of each other (standard plate test and preincubation assay).
Besides, the results of the negative as well as the positive controls performed in parallel corroborated the validity of this study, since the values fulfilled the acceptance criteria of this study. In this study with and without S9 mix, the number of revertant colonies in the negative controls was within or marginally below the range of the historical negative control data for each tester strain. In addition, the positive control substances both with and without S9 mix induced a significant increase in the number of revertant colonies within the range of the historical positive control data.
Thus, under the experimental conditions chosen here, it is concluded that the test substance is not mutagenic in the bacterial reverse mutation test in the absence and the presence of metabolic activation.

Justification for classification or non-classification

Based on the study and taking into account the provisions laid down in Regulation (EC) No 1272/2008 as amended for the ninth time in Regulation (EU) No 2016/1179, the test substance does not need to be classified as mutagenic.