Benzenemethanimine, .alpha.-phenyl-

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test substance showed no mutagenic effect in a Ames tests with 4 strains of Salmonella typhimurium with and without metabolic activation (OECD TG 471, GLP).

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:

guideline study with acceptable restrictions

Remarks:

Study comparable to OECD Guideline 471 under GLP conditions with acceptable restrictions (E. coli WP2 or S. typhimurium TA102 missing)

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

E. coli WP2 or S. typhimurium TA102 missing

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

- Name of test material (as cited in study report): BPI (Benzophenone imine)
- Analytical purity: 94.8 %
- Lot/batch No.: 11/94, Container 13
- Physical state: light yellow liquid
- Storage condition of test material: refrigerator
- Stability under test conditions: The stability of the test substance throughout the study period has been proven by reanalysis. The stability of the test substance in the vehicle DMSO over a period of 4 hours and in water over a period of 1 hour was verified analytically.

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9-Mix prepared from Aroclor 1254-induced Sprague Dawley rat livers

Test concentrations with justification for top dose:

20 - 5000 µg/plate (standard plate test); 0.8 - 500 µg/plate (preincubation test)

Vehicle / solvent:

- Vehicle used: DMSO

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Remarks:

with S9: 2.5 µg 2-aminoanthracene for all strains; without S9: 5 µg N-methyl-N'-nitro-N-nitrosoguanidine for strains TA100 and TA1535, 10 µg 4-nitro-o-phenylendiamine for strain TA98, 100 µg 9-aminoacridine chloride monohydrate for strain TA 1537.

Positive control substance:

9-aminoacridine

N-ethyl-N-nitro-N-nitrosoguanidine

other: with S9: 2.5 µg 2-aminoanthracene for all strains; without S9: 10 µg 4-nitro-o-phenylendiamine for strain TA98.

Details on test system and experimental conditions:

Standard Plate Test
Test tubes containing 2 ml soft agar kept in a water bath at 45°C, and remaining components added in the following order:
0.1 ml test solution or vehicle
0.1 ml bacterial suspension
0.5 ml S-9 mix (in tests with metabolic activation) or 0 .5 ml phosphate buffer (in tests without metabolic activation).
After mixing, the samples are poured onto Vogel-Bonner agar plates.

Preincubation assay
0.1 ml test solution or vehicle, 0.1 ml bacterial suspension and 0 .5 ml S-9 mix are incubated at 37°C for the duration of 20 minutes. Subsequently, 2 ml of soft agar is added and, after mixing, the samples are poured onto the Vogel-Bonner agar plates.

Both Tests
In each experiment 3 test plates per dose or per control used; after incubation at 37°C for 48 hours in the dark, the bacterial colonies ( his+ revertants) are counted.

The titer was determined and in regularly measurements the strain characteristics were checked. Sterility control was performed.

Evaluation criteria:

To be chracterized as positive, a substance has to fulfill the following requirements:
- doubling of the spontaneous mutation rate (control)
- dose-response relationship
- reproducibility of the results

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

A bacteriotoxic effect (reduced his background growth, decrease in the number of his+ revertants) was observed at doses > 500 µg/plate (standard plate test) or from about 20 - 100 µg/plate onward (preincubation test).

Vehicle 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:

A bacteriotoxic effect (reduced his background growth, decrease in the number of his+ revertants) was observed at doses > 500 µg/plate (standard plate test) or from about 20 - 100 µg/plate onward (preincubation test).

Vehicle 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:

A bacteriotoxic effect (reduced his background growth, decrease in the number of his+ revertants) was observed at doses > 500 µg/plate (standard plate test) or from about 20 - 100 µg/plate onward (preincubation test).

Vehicle 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:

A bacteriotoxic effect (reduced his background growth, decrease in the number of his+ revertants) was observed at doses > 500 µg/plate (standard plate test) or from about 20 - 100 µg/plate onward (preincubation test).

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitation of the test substance was found

Conclusions:

The test substance is not mutagenic in the Ames test with and without metabolic activation under the experimental conditions chosen.

Executive summary:

The test substance was tested for its mutagenic potential based on the ability to induce back mutations in selected loci in several strains of Salmonella typhimurium in the Ame.The study was conducted under GLP conditions and is comparable to OECD Guideline 471 with acceptable restrictions (E. coli strain WP2 or S. typhimurium strain TA102 missing).

Strains: TA 1535, TA 100, TA 1537, TA 98

Dose range: 20.0 µg -5,000 µg/plate (SPT), 0.8 µg -500 µg/plate (PIT)

Test conditions: Standard plate test (SPT) and preincubation test (PIT) both with and without metabolic activation (Aroclor induced rat liver 5-9mix).

Solubility: No precipitation of the test substance was found.

Toxicity: A bacteriotoxic effect was observedat doses > 500 pg/plate (SPT) or from about 20 µg -100 µg/plate onward (PIT).

Mutagenicity: An increase in the number of his+ revertants was not observed both in the standard plate test and in the preincubation test either without S-9 mix or after the addition of a metabolizing system.

Conclusion: According to the results of the present study, the test substance is not mutagenic in the Ames test under the experimental conditions chosen here.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

The test substance was tested for its mutagenic potential based on the ability to induce back mutations in selected loci in several strains of Salmonella typhimurium in the Ame.The study was conducted under GLP conditions and is comparable to OECD Guideline 471 with acceptable restrictions (E. coli strain WP2 or S. typhimurium strain TA102 missing).

Strains: TA 1535, TA 100, TA 1537, TA 98

Dose range: 20.0 µg -5,000 µg/plate (SPT),0.8 µg -500 µg/plate (PIT)

Test conditions: Standard plate test (SPT) and preincubation test (PIT) both with and without metabolic activation (Aroclor induced rat liver 5-9mix).

Solubility: No precipitation of the test substance was found.

Toxicity: A bacteriotoxic effect was observedat doses > 500 pg/plate (SPT) or from about 20 µg -100 µg/plate onward (PIT).

Mutagenicity: An increase in the number of his+ revertants was not observed both in the standard plate test and in the preincubation test either without S-9 mix or after the addition of a metabolizing system.

Conclusion: According to the results of the present study, the test substance is not mutagenic in the Ames test under the experimental conditions chosen here.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. No indication of genotoxicity was observed in the Ames test (OECD 471, GLP). As a result, the substance is not classified for genotoxicity under Regulation (EC) No. 1272/2008, as amended for the thirteenth time in Regulation (EC) No. 2018/1480.