4-methyl-4-phenylpentan-2-one

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames test (OECD 471): negative in S. typhimurium TA 1535, TA 1537, TA 98 and TA 100 and E. coli WP2 uvr A with and without metabolic activation

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:

28 Feb - 31 Mar 2017

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:

adopted 21 Jul 1997

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Commission Regulation (EC) No. 440/2008 B13/14, dated 30 May 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hess. Ministerium für Umwelt, Klimaschutz, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany

Type of assay:

bacterial reverse mutation assay

Target gene:

his opron (for S. typhimurium strains)
trp operon (for E.coli strain)

Species / strain / cell type:

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

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with Phenobarbital/β-naphthoflavone

Test concentrations with justification for top dose:

Pre-experiment: 3, 10, 33, 100, 333, 1000, 2500 and 5000 µg/plate with and without metabolic activation
The pre-experiment is reported as Experiment 1.

Experiment 2:
3, 10, 33, 100, 333, 1000, 2500 and 5000 µg/plate with and without metabolic activation for TA 100 and WP2 uvrA
33, 100, 333, 1000, 2500 and 5000 µg/plate with and without metabolic activation for remaining strains

Vehicle / solvent:

- Solvent used: DMSO
- Justification for choice of solvent: The solvent was chosen because of its solubility properties and its relative nontoxicity to the bacteria.

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive control substance:

sodium azide

methylmethanesulfonate

other: 4-nitro-o-phenylene-diamine (4-NOPD), 2-aminoanthracene (2-AA)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation) (Experiment 1); preincubation (Experiment 2)

DURATION
- Preincubation period: 1 h
- Exposure duration: at least 48 h

NUMBER OF REPLICATIONS: 3 replications each in 2 independent experiment

DETERMINATION OF CYTOTOXICITY
- Method: reduction in the number of spontaneous revertants or a clearing of the bacterial background lawn

Evaluation criteria:

A test substance is considered as mutagenic if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100 and WP2 uvr A) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
A dose-dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
A dose-dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant.

Statistics:

Mean values and standard deviations were calculated.

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Exp 2: +/- S9: at and above 1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Exp 1: +/- S9: at 5000 µg/plate; Exp 2: +/- S9: at and above 1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Exp 1: + S9: at 5000 µg/plate; Exp 2: +/- S9: at and above 1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Exp 1: +/- S9: at and above 1000 µg/plate; Exp 2: +/- S9: at and above 1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Exp 1: +S9: at and above 2500 µg/plate; Exp 2: +/- S9: at and above 1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: The test substance precipitated in the overlay agar in the test tubes at 5000 μg/plate without S9 mix and from 2500 to 5000 μg/plate with S9 mix in both experiments. Precipitation of the test substance in the overlay agar on the incubated agar plates was observed only in Experiment 1 at 5000 μg/plate with S9 mix. The undissolved particles had no influence on the data recording.

RANGE-FINDING/SCREENING STUDIES: The pre-experiment is reported as Experiment 1 (all strains were tested in the pre-experiment).

ADDITIONAL INFORMATION ON CYTOTOXICITY: Toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in all strains. Reduced background growth was not observed in Experiment 1 at any test substance concentration with or without metabolic activation. In Experiment 2 reduced background growth was observed with and without metabolic activation in all strains at and above 1000 µg/plate.

Table 1. Test results of Experiment 1 (plate incorporation).

	Number of revertant colonies (mean of 3 plates±SD)
Without S9
Test substance (µg/plate)	TA 1535	TA 1537			TA 98	TA 100		WP2 uvr A
DMSO	11±4	8±3			23±7	205±6		38±0
Untreated	15±4	10±3			23±4	206±14		47±11
3	9±2	8±3			24±5	190±28		44±11
10	8±1	8±2			20±6	182±20		38±6
33	13±6	9±2			25±7	195±16		42±6
100	9±2	8±2			27±5	205±13		43±2
333	14±3	11±1			27±3	209±10		44±5
1000	14±3	9±2			22±3	82±12		29±10
2500	12±3	8±3			22±8	88±10		30±8
5000	15±3	3±2			18±2	44±11		29±4
Positive Control	NaN3	4-NOPD			4-NOPD	NaN3		MMS
Dose (µg/plate)	10	50			10	10		2
Number of revertant colonies/plate	1457±44	90±11			443±15	2147±136		930±35
With S9
Test substance (µg/plate)	TA 1535		TA 1537		TA 98	TA 100	WP2 uvr A
DMSO	13±4		11±1		45±7	152±19	64±4
Untreated	8±3		10±3		44±7	196±11	61±4
3	14±4		13±2		39±3	143±28	57±12
10	14±3		16±3		35±3	149±10	55±6
33	12±3		13±6		37±10	164±15	52±8
100	11±2		8±2		43±3	175±4	50±11
333	14±4		13±1		28±8	168±18	51±7
1000	10±4		13±6		32±4	52±1	34±2
2500	14±2		11±1		23±6	45±11	27±8
5000	6±2 p,m		1±1 p		0±0	0±1 p	8±2 p,m
Positive Control	2-AA		2-AA		2-AA	2-AA	2-AA
Dose (µg/plate)	2.5		2.5		2.5	2.5	10
Number of revertant colonies/plate	439±14		146±10		4202±220	3893±20	339±11
NaN3: sodium azide 2AA: 2-Aminoanthracene 4-NOPD: 4-nitro-o-phenylene-diamine MMS: methyl methane silfonate				p: precipitate m: manual count

Table 2. Test results of Experiment 2 (preincubation).

	Number of revertant colonies (mean of 3 plates±SD)
Without S9
Test substance (µg/plate)	TA 1535	TA 1537			TA 98	TA 100		WP2 uvr A
DMSO	12±2	9±4			29±3	155±15		42±12
Untreated	9±3	10±4			30±3	178±21		49±5
3						158±7		54±14
10						146±3		42±5
33	10±2	9±5			31±2	148±21		44±4
100	12±4	7±2			30±2	149±7		43±8
333	16±5	9±2			25±5	126±4		41±10
1000	9±0 r	3±1 r			8±1 r,m	75±15 r		27±6 r
2500	13±2 r	4±1 r			11±2 r,m	46±3 r		15±2 r,m
5000	15±1 r	0±0 r			14±2 r	3±1 r,m		3±1 r
Positive Control	NaN3	4-NOPD			4-NOPD	NaN3		MMS
Dose (µg/plate)	10	50			10	10		2
Number of revertant colonies/plate	1246±373	110±10			388±53	2206±129		841±37
With S9
Test substance (µg/plate)	TA 1535		TA 1537		TA 98	TA 100	WP2 uvr A
DMSO	14±3		14±3		34±3	122±10	50±12
Untreated	15±4		19±3		43±7	166±9	52±7
3						125±15	49±13
10						126±11	55±5
33	11±3		15±5		39±8	126±12	50±12
100	11±4		11±1		34±5	120±11	55±5
333	11±1		12±3		34±7	78±9	48±10
1000	19±2 r		6±1 r		23±7 r	33±0 r	27±5 r
2500	1±0 r		2±2 r		1±1 r	4±1 r	34±12 r
5000	0±0 r		0±1 r		1±1 r	1±1 r	3±1 r
Positive Control	2-AA		2-AA		2-AA	2-AA	2-AA
Dose (µg/plate)	2.5		2.5		2.5	2.5	10
Number of revertant colonies/plate	337±15		226±41		4213±411	4425±143	511±36
NaN3: sodium azide 2AA: 2-Aminoanthracene 4-NOPD: 4-nitro-o-phenylene-diamine MMS: methyl methane silfonate				r: reduced background growth m: manual count

Conclusions:

Under the conditions of the conducted test the substance was not mutagenic in any of the five strains (TA 1535, TA 1537, TA 98, TA 100 and WP2 uvrA) tested with and without metabolic activation up to 5000 µg/plate.

Executive summary:

A bacterial gene mutation assay with the test substance was performed in accordance with OECD Guideline 471 and in compliance with GLP (2017). In this study the substance was not mutagenic in any of the five strains (TA 1535, TA 1537, TA 98, TA 100 and WP2 uvr A) tested with and without metabolic activation up to and including 5000 µg/plate.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

A bacterial gene mutation assay with the test substance was performed in accordance with OECD Guideline 471 and in compliance with GLP (2017). In this study the substance was not mutagenic in any of the five strains (TA 1535, TA 1537, TA 98, TA 100 and WP2 uvr A) tested with and without metabolic activation up to and including 5000 µg/plate.

Justification for classification or non-classification

The available data on genetic toxicity meet criteria for data requirements of Annex VII but is not sufficient for classification or non-classification according to Regulation (EC) 1272/2008.