Benzyl phenylacetate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Gene mutation toxicity study was performed to determine the mutagenic nature of benzyl phenylacetate using preincubation assay. The study was performed using Salmonella typhimurium strain TA97, TA98, TA1535 and TA100 with and without 10% and 30% rat liver and hamster liver S9 metabolic activation system. The study was performed at dose level of 0, 0.3, 1, 3, 10, 33, 66, 100, 333, 1000, 3333 or 10000 µg/plate for TA100 and TA98 and at a dose level of 0, 0.3, 1, 3, 10, 33, 100, 333, 1000, 3333 or 10000 µg/plate for TA1535 and TA97 with DMSO as the solvent. The plates were observed for a dose dependent increase in the number of revertants/plate. Benzyl phenylacetate did not induce a a dose dependent increase in the number of revertants per plate and hence exhibited no mutagenic activity under the given test conditions.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from CEBS (Chemical Effect in Biological System) report

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Gene mutation study by the preincubation assay was performed to determine the mutagenic nature of benzyl phenylacetate

GLP compliance:

no

Type of assay:

bacterial gene mutation assay

Specific details on test material used for the study:

- Name of test material: Benzyl phenylacetate
- Molecular formula: C15H14O2
- Molecular weight: 226.2736 g/mol
- Substance type: Organic
- Physical state: No data
- Purity: No data
- Impurities (identity and concentrations): No data

Target gene:

Histidine

Species / strain / cell type:

S. typhimurium, other: TA97, TA98, TA100 and TA1535

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

with and without

Metabolic activation system:

10% and 30% rat liver and hamster liver S9 fraction

Test concentrations with justification for top dose:

TA100 and TA98: 0, 0.3, 1, 3, 10, 33, 66, 100, 333, 1000, 3333 or 10000 µg/plate
TA 1535 and TA1537: 0, 0.3, 1, 3, 10, 33, 100, 333, 1000, 3333 or 10000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The chemical was solubel in DMSO

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

not specified

Details on test system and experimental conditions:

METHOD OF APPLICATION : Pre incubation

DURATION
No data

SELECTION AGENT (mutation assays):No data
SPINDLE INHIBITOR (cytogenetic assays):No data
STAIN (for cytogenetic assays):No data

NUMBER OF REPLICATIONS: No data

NUMBER OF CELLS EVALUATED:
No data

METHOD OF APPLICATION: preincubation
- Cell density at seeding (if applicable): No data

DURATION
- Preincubation period: No data
- Exposure duration: No data
- Expression time (cells in growth medium): No data
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data

SPINDLE INHIBITOR (cytogenetic assays): v

STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: No data

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: No data

NUMBER OF CELLS EVALUATED: No data

NUMBER OF METAPHASE SPREADS ANALYSED PER DOSE (if in vitro cytogenicity study in mammalian cells): No data

CRITERIA FOR MICRONUCLEUS IDENTIFICATION: No data

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data
- Any supplementary information relevant to cytotoxicity: Yes, cytotoxicity was noted

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Methods, such as kinetochore antibody binding, to characterize whether micronuclei contain whole or fragmented chromosomes (if applicable): No data

- OTHER:

Rationale for test conditions:

No data

Evaluation criteria:

The plates were observed for a dose dependent increase in the number of revertants/plate

Statistics:

Mean ± standard error of mean

Species / strain:

S. typhimurium, other: TA97, TA98, TA100 and TA1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Remarks:

Slight toxicity was noted

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

No data

Strain: TA100

Dose	No Activation (Negative)		No Activation (Negative)		30% RLI (Negative)		30% HLI (Negative)		10% RLI (Negative)		10% HLI (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
µg/plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	141	7.3	106	3.7	161	5.1	165	4.1	145	7	117	8.2
0.3			101	2.3
1	154	4.3	104	4
3	154	10.8	97	6.9
10	150	14.6	109	8
33	152	835	89	1.7
66	58 s	4.8
100					157	1.5	190	11.7	130	9.9	123	3.6
333					180	7.1	161	12.8	128	3.5	130	10.1
1000					163	1.7	190	9.7	122	5.8	118	10.1
3333					173	4.7	148	20.2	106	12.4	116	3.5
10000					153	7.9	186	17.3	104	13.8	105	5.9
Positive Control	466	15.3	7.4	58.5	443	42.6	638	10.9	422	16.5	587	43.3

 

Strain :TA1535

Dose	No Activation (Negative)		No Activation (Negative)		30% RLI (Negative)		30% HLI (Negative)		10% RLI (Negative)		10% HLI (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
µg/plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	12 12	1.9	12	1.5	12	0.3	13	2.1	13	2	14	2.9
0.3	12	1.2	8	0.9
1	9	0.9	9	0.7
3	10	1.2	7	0.9
10	10	2.1	10	2.3
33	9	1	10	1.9
100					19.3	0.3	12	0.9	13	2.3	12	0.3
333					15	2.3	8	1.5	13	3.7	10	2.6
1000					12	0.3	4	1.2	8	2.2	12	1.7
3333					9	0.6	8	1.5	10	1.5	9	1.5
10000					8	1.2	7	0.7	8	2	10	2.2
Positive Control	585	19.9	635	37.5	106	10.8	256	24.9	71	8.1	61	0.9

 

Strain :TA97

Dose	No Activation (Negative)		No Activation (Negative)		30% RLI (Negative)		30% HLI (Negative)		10% RLI (Negative)		10% HLI (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
µg/plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	203	6.2	199	0.9	231	8	218	10.9	188	5.5	168	10
0.3	207	4.1	179	3.7
1	223	8.5	125	5
3	199	7.5	182	10.4
10	189	4	176	7.2
33	191	4.8	185	13.7
100					237	11.7	227	2.6	190	5.6	187	8.6
333					242 c	15.5	211	5.9	220	9.7	189	15.8
1000					243	6.6	199	11.6	187	1.8	164	12.7
3333					254	15.7	204	16.3	148	10.4	146	4.3
10000					190	2.3	185	14.6	162	13.3	128	5
Positive Control	554	8.3	470	27.7	378	19.7	426	12.7	282	8.6	342	48.5

 

Strain :TA98

Dose	No Activation (Negative)		No Activation (Negative)		30% RLI (Negative)		30% HLI (Negative)		10% RLI (Negative)		10% HLI (Negative)		10% RLI (Negative)		10% HLI (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
µg/plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	22	3.1	21	3.7	23	3.1	20	2.6	27	3.2	20	2.4	23	0.3	22	5.8
0.3			17	0
1	24	2.5	20	4.4
3	27	1.8	12	3.2
10	22	1.7	19	2.8							21	2.3			16	2.1
33	15	1.5	17	0.6							19	1.5			15	1.5
66	4s	1.7
100					23	44	22	2.6	18	1.5	14	1.2	17	0.6	18	1
333					24	2.7	24	4.8	19	4.3	8	1.9	16	0.6	12	2.3
100					16 2	2	22	4.3	13	2.2	5s	0.9	11	1.2	11	0.9
3333					15	2.9	18	1.5	9s	2.3			7s	0.9
1000					12 s	1.9	10	2.2	5s	0.9			4s	1
Positive Control	399	23.8	894	16.8	129	3.8	345	9.1	332	30.3	341	15	473	18.5	584	7.8

RLI = induced male Sprague Dawley rat liver S9

HLI = induced male Syrian hamster liver S9

s = Slight Toxicity; p = Precipitate; x = Slight Toxicity and Precipitate; t = Toxic; c = Contamination

Conclusions:

Benzyl phenylacetate did not induce a a dose dependent increase in the number of revertants per plate in Salmonella typhimurium strain TA97, TA98, TA1535 and TA100 with and without 10% and 30% rat liver and hamster liver S9 metabolic activation system and hence exhibited no mutagenic activity under the given test conditions.

Executive summary:

Gene mutation toxicity study was performed to determine the mutagenic nature of benzyl phenylacetate using preincubation assay. The study was performed using Salmonella typhimurium strain TA97, TA98, TA1535 and TA100 with and without 10% and 30% rat liver and hamster liver S9 metabolic activation system. The study was performed at dose level of 0, 0.3, 1, 3, 10, 33, 66, 100, 333, 1000, 3333 or 10000 µg/plate for TA100 and TA98 and at a dose level of

0, 0.3, 1, 3, 10, 33, 100, 333, 1000, 3333 or 10000 µg/plate for TA1535 and TA97 with DMSO as the solvent. The plates were observed for a dose dependent increase in the number of revertants/plate. Benzyl phenylacetate did not induce a dose dependent increase in the number of revertants per plate and hence exhibited no mutagenic activity under the given test conditions.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Gene mutation in vitro:

Various data available for the target chemical Benzyl phenylacetate was reviewed to determine its mutagenic nature. The summary is as mentioned below:

Gene mutation toxicity study was performed to determine the mutagenic nature of benzyl phenylacetate using preincubation assay (National Institute of Environmetal Health Sciences, 2015). The study was performed using Salmonella typhimurium strain TA97, TA98, TA1535 and TA100 with and without 10% and 30% rat liver and hamster liver S9 metabolic activation system. The study was performed at dose level of 0, 0.3, 1, 3, 10, 33, 66, 100, 333, 1000, 3333 or 10000 µg/plate for TA100 and TA98 and at a dose level of 0, 0.3, 1, 3, 10, 33, 100, 333, 1000, 3333 or 10000 µg/plate for TA1535 and TA97 with DMSO as the solvent. The plates were observed for a dose dependent increase in the number of revertants/plate. Benzyl phenylacetate did not induce a dose dependent increase in the number of revertants per plate and hence exhibited no mutagenic activity under the given test conditions.

In another study by Zeiger and Margolin (2000), gene mutation toxicity study was performed to determine the mutagenic nature of Benzyl phenylacetate. The study was performed as per the preincubation modification of the Salmonella/mammalian microsome mutagenicity (Ames) test. The chemicals were tested in a preincubation procedure in strains TA98 and TA100 without metabolic activation and with activation provided by Aroclor induced rat and hamster liver homogenates (S9). If a positive response was seen in one of these two strains, the strain/metabolic activation combination producing that response was repeated, and no further testing was performed. If no positive responses were seen, the chemical was tested in strains TA97 and TA1535. The plates were observed for a dose dependent increase in the number of revertants/plate. The combination of a questionable (?) and negative (-) response was considered negative (-); the combination of a weakly positive (+w) and negative response was considered questionable (?). Benzyl phenylacetate did not induce a dose dependent increase in the number of revertants in Salmonella typhimurium TA98, TA100, TA97 and TA1535 in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Renne et al (2006) performed gene mutation toxicity studu using cigarette smoke. The mutagenicity of total particulate matter (TPM) in cigarettes was investigated using an Ames assay protocol that conformed to OECD Guideline 471. For this purpose, prototype cigarettes containing a mixture of ingredients, reference cigarettes without these ingredients, and 2R4F cigarettes (a standard reference cigarette). The concentration of benzyl phenylacetate in the test cigarrette was 2.6 ppm. Each sample was tested with and without S9 metabolic activation in five strains ofSalmonella typhimuriumTA98, TA100, TA102, TA1535, and TA1537. Evaluation of the Ames assay data was carried out as mutagenic response, taking into consideration the reproducibly dose-related increase in number of revertants, even if the increase was less than two-fold. The results of Ames assay on test cigarette with and without metabolic activation. TA100, TA98, and TA1537 strains showed a positive response only with metabolic activation. No response was observed in TA 102 or TA1535. No sporadic responses in revertants were recorded. The highest sensitivity and specificity of the mutagenic response were observed using TA98 with metabolic activation. From the comparison of the data obtained for the test and reference cigarettes, it was concluded that the addition of ingredients did not result in a positive mutagenic response in any of the strains under the conditions already described. Benzyl phenylacetate can be considered to be non mutagenic at a test concentration of 2.6 ppm under the test conditions as it did not induce a reproducible dose dependent increase in the number of revertants.

Based on the data available for the target chemical, Benzyl phenylacetate does not exhibit gene mutation ability in vitro. Hence, the target chemical is not likely to classify as a gene mutant in vitro as per the criteria mentioned in CLP regulation.

Justification for classification or non-classification

Based on the data available for the target chemical, Benzyl phenylacetate does not exhibit gene mutation ability in vitro. Hence, the target chemical is not likely to classify as a gene mutant in vitro as per the criteria mentioned in CLP regulation.