Reaction mass of 4-methylene-2-phenyltetrahydro-2H-pyran and 4-methyl-2-phenyl-3,6-dihydro-2H-pyran

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Under the test conditions, Reaction mass of 3,6-dihydro-4-methyl-2-phenyl-2H-pyran and tetrahydro-4-methylene-2-phenyl-2H-pyran is not considered as mutagenic in S. typhimurium (TA1535, TA1537, TA1538, TA98 and TA100) strains. The QSAR on TA102 in the presence and absence of S9-mix are also both predicted to be negatibe with strong confidence.

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

Study period:

22 December 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP study conducted similarly to OECD 471 Guideline with deviations such as purity of test item, evaluation and acceptance criteria and individual plate counts not reported

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

no GLP statement, purity of test item, evaluation and acceptance criteria and individual plate counts not reported

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Target gene:

None

Species / strain / cell type:

S. typhimurium, other: TA 1535, TA 1537, TA 1538, TA 98 and TA 100

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

other: strains were checked for numbers of spontaneous revertants, histidine requirement and sensitivity to ampicillin, crystal violet and UV radiation.

Metabolic activation:

with and without

Metabolic activation system:

10 % S9 mix; S9 fraction prepared from liver homogenates of male Wistar rats intraperitoneally induced with Aroclor 1254 (500 mg/kg bw) in soya bean oil

Test concentrations with justification for top dose:

Mutagenicity test:
- 0.0012, 0.004, 0.011, 0.033 and 0.1 mg/plate (in 0.1 mL methanol per plate), with and without S9 mix in TA 1535, TA 1537, TA 1538, TA 98 and TA 100 strains

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Methanol
- Test substance preparation: Appropriate dilutions of the test material in methanol were prepared immediately before use.

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

methanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: sodium azide (0.5 µg/plate (in 0.1 mL water)) for strains TA 1535 and TA 100 ; hycanthone methanesulphonate (12.5 µg/plate (in 0.1 mL water)) for strains TA 1537, TA 1538 and TA 98

Remarks:

without S-9 mix

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

methanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene (0.5 µg/plate (in 0.1 mL DMSO) for all strains

Remarks:

with S-9 mix

Details on test system and experimental conditions:

SOURCE OF TEST SYSTEM: - All Salmonella typhimurium strains received from Dr. B.N. Ames, Berkeley, California, U.S.A.

METHOD OF APPLICATION: In agar (plate incorporation)

DURATION
- Incubation period: Plates were incubated at 37 °C for three days

NUMBER OF REPLICATIONS:
- 3 plates/dose for treatment, vehicle and positive controls

DETERMINATION OF CYTOTOXICITY
- Method: Toxicity was determined based on background lawn of bacterial growth.

OTHER:
- Colony counting: After incubation, the colonies (revertants which are histidine independent) were counted and the background lawn of bacterial growth was examined microscopically.

Evaluation criteria:

None

Statistics:

None

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 1538, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

CYTOTOXICITY:
- In preliminary toxicity test, dose-levels of 10 mg/plate and higher appeared to abolish completely all bacterial growth, while at a level of 1 mg/plate the test material was still toxic for the bacteria as revealed by a less dense background lawn of bacterial growth and a decrease in the numbers of his+ revertants.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

See the attached document for information on tables of results

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

Under the test conditions, Reaction mass of 3,6-dihydro-4-methyl-2-phenyl-2H-pyran and tetrahydro-4-methylene-2-phenyl-2H-pyran is not considered as mutagenic in S. typhimurium (TA1535, TA1537, TA1538, TA98 and TA100) strains.

Executive summary:

In a reverse gene mutation assay in bacteria, performed similarly to the OECD Guideline 471, strains of Salmonella typhimurium (TA1535, TA1537, TA1538, TA98 and TA100) were exposed to Reaction mass of 3,6-dihydro-4-methyl-2-phenyl-2H-pyran and tetrahydro-4-methylene-2-phenyl-2H-pyran at the concentrations of 0.0012, 0.004, 0.011, 0.033 and 0.1 mg/plate (in 0.1 mL methanol per plate), with and without S9 mix. Metabolic activation system used in this test 10 % S9 mix; S9 fraction prepared from liver homogenates of male Wistar rats intraperitoneally induced with Aroclor 1254 (500 mg/kg bw) in soya bean oil. Vehicle and positive control groups were also included in mutagenicity tests.

 

In preliminary toxicity test, dose-levels of 10 mg/plate and higher appeared to abolish completely all bacterial growth, while at a level of 1 mg/plate the test material was still toxic for the bacteria as revealed by a less dense background lawn of bacterial growth and a decrease in the numbers of his+ revertants. Incorporation of the test product with the bacteria at levels up to 0.1 mg/plate did not increase the numbers of his+ revertants with any of the five tester strains, either in the presence or in the absence of S-9 mix. There were no signs that chemical toxicity interfered with the mutagenicity testing: the background lawn of bacterial growth in control and test plates was comparable. The positive and vehicle controls induced the appropriate responses in the corresponding strains indicating the validity of the study.

 

Under the test conditions, Reaction mass of 3,6-dihydro-4-methyl-2-phenyl-2H-pyran and tetrahydro-4-methylene-2-phenyl-2H-pyran is not considered as mutagenic in bacteria.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

Justification for selection of genetic toxicity endpoint
A reliable in vivo study

Justification for classification or non-classification

Based on a negative results in the reliable bacterial reverse mutation assay five strains of Salmonella typhimurium, classification under EU DSD or CLP regulations as a mutgen is not required.