3,9-bis[2,4-bis(1-methyl-1-phenylethyl)phenoxy]-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane;bis(2,4-dicumylphenyl) neopentyl diphosphite

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Gene mutation (Bacterial Reverse Mutation Assay/Ames test): the substance was not mutagenic in the strains Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537 & Escherichia coli WP2, WP2uvrA in the presence and absence of Aroclor 1254-induced rat liver S9 metabolic activation. (Directive 92/69 EEC, B.14/GLP).

In vitro cytogenicity (chromosome aberration) study in mammalian cells: the substance did not induce any induce structural and/or numerical chromosomal damage in CHO cells in the presence or absence of Aroclor 1254-induced rat liver S9 metabolic activation (Directive 92/69/EEC, B.10/GLP).

Gene mutation (in vitro mammalian cell gene mutation): there was no evidence of induced mutant colonies over background in CHO cells exposed to the substance in the presence or absence of Aroclor 1254-induced rat liver S9 metabolic activation (Directive 87/302 EEC, L.133 p.61-63/GLP).

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP Compliance, not OECD guideline

Qualifier:

according to guideline

Guideline:

other: Directive 92/69 EEC, B.14 Ames-test

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

bacteria, other: Salmonella typhimurium & Escherichia coli TA 98, TA 100, TA 1535, TA 1537 & WP2, WP2uvrA

Metabolic activation system:

S9 liver fractions of rats treated with AROCLOR 1254

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 33 ... 5000 μg/plate
Concentration range in the main test (without metabolic activation): 33 ... 5000 μg/plate

Vehicle / solvent:

Solvent: DMSO

Species / strain:

other: as specified above

Remarks:

Preliminary test

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 μg/plate)

Species / strain:

other: as specified above

Remarks:

Preliminary test

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 μg/plate)

Species / strain:

other: as specified above

Remarks:

Main test

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 μg/plate)

Species / strain:

other: as specified above

Remarks:

Main test

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 μg/plate)

Additional information on results:

Observations:
Due to the poor solubility, the test substance was tested as suspension.
The undissolved particles had no influence on the data recording.

Conclusions:

Interpretation of results:
negative with metabolic activation
negative without metabolic activation

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP Compliance, not OECD guideline

Qualifier:

according to guideline

Guideline:

other: Directive 92/69 EEC, B.10

GLP compliance:

yes

Type of assay:

other: in vitro mammalian cytogenicity (B10)

Species / strain / cell type:

mammalian cell line, other: CHO-cells of the Chinese hamster

Metabolic activation system:

S9 liver fractions of rats treated with AROCLOR 1254

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 3 - 5000 μg/ml
Concentration range in the main test (with metabolic activation): 1 - 5000 μg/ml
Concentration range in the main test (without metabolic activation): 3 - 5000 μg/ml
Concentration range in the main test (without metabolic activation): 1 - 1000 μg/ml

Vehicle / solvent:

DMSO

Details on test system and experimental conditions:

Exposure period (with metabolic activation): 4 hours
Exposure period (without metabolic activation): 22 hours
Fixation time: 22 and 30 hours

Species / strain:

other: as specified above

Remarks:

Preliminary test

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(≥3000 μg/ml)

Species / strain:

other: as specified above

Remarks:

Preliminary test

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(≥300 μg/ml)

Species / strain:

other: as specified above

Remarks:

Main test

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(≥3 μg/ml)

Species / strain:

other: as specified above

Remarks:

Main test

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(≥1000 μg/ml)

Additional information on results:

Observations:
Precipitation was observed at concentrations ≥30μg/ml (with and without S9-mix).

Conclusions:

Interpretation of results:
negative with metabolic activation
negative without metabolic activation

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP Compliance, not OECD guideline

Qualifier:

according to guideline

Guideline:

other: Directive 87/302 EEC, L.133 p.61-63

GLP compliance:

yes

Type of assay:

other: in vitro mammalian cell gene mutation (B17)

Species / strain / cell type:

mammalian cell line, other: - CHO Cells of Chinese hamster

Metabolic activation system:

S9 liver fractions of rats treated with AROCLOR 1254

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 10 - 3000 μg/ml
Concentration range in the main test (with metabolic activation): 10 - 5000 μg/ml
Concentration range in the main test (without metabolic activation): 10 - 3000 μg/ml
Concentration range in the main test (without metabolic activation): 10 - 300 μg/ml

Vehicle / solvent:

DMSO

Details on test system and experimental conditions:

Exposure period (with metabolic activation): 4 hours
Exposure period (without metabolic activation): 4 hours
Expression time: 7 days
Selection time:- 9 days
Fixation time:- Not applicable

Species / strain:

other: as specified above

Remarks:

Preliminary test

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(5000 μg/ml)

Species / strain:

other: as specified above

Remarks:

Preliminary test

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(3000 μg/ml)

Species / strain:

other: as specified above

Remarks:

Main test

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(5000 μg/ml)

Species / strain:

other: as specified above

Remarks:

Main test

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(≥300 μg/ml)

Additional information on results:

Observations:
- Precipitation was observed at concentrations ≥300 μg/ml (with and without S9-mix).

Conclusions:

Interpretation of results:
negative with metabolic activation
negative without metabolic activation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Gene mutation (Bacterial Reverse Mutation Assay/Ames test):

There is one gene mutation study (Bacterial Reverse Mutation Assay/Ames test) available.

In a reverse gene mutation assay in bacteria (Directive 92/69 EEC, B.14/GLP), strains of Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537 & Escherichia coli WP2, WP2uvrA were exposed to 3,9-bis[2,4-bis(1-methyl-1-phenylethyl)phenoxy]-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane in DMSO at concentrations of 33-5000µg/plate in the presence and absence of Aroclor 1254-induced rat liver S9 metabolic activation. Due to the poor solubility, the test substance was tested as suspension. The undissolved particles had no influence on the data recording. Cytotoxicity was noted in the preliminary and main tests (>5000 ug/plate). The substance was negative with and without metabolic activation.

In vitro cytogenicity (chromosome aberration) study in mammalian cells:

There is one in vitro cytogenicity (chromosome aberration) study in mammalian cells available.

In an in vitro cytogenicity (chromosome aberration) study (Directive 92/69/EEC, B.10/GLP), CHO cells were exposed to 3,9-bis[2,4-bis(1-methyl-1-phenylethyl)phenoxy]-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane in DMSO at concentrations from 1 - 5000µg/ml with (for 4 hrs) and without (22 hrs) Aroclor 1254-induced rat liver S9 metabolic activation. Cytotoxicity was observed at ≥3000 µg/ml with metabolic activation and ≥300 µg/ml without metabolic activation in the preliminary tests. Cytotoxicity was observed at ≥3 µg/ml with metabolic activation and ≥1000 µg/ml without metabolic activation in the main tests. Precipitation was observed at ≥30 μg/ml with and without metabolic activation.The substance did not induce structural and/or numerical chromosomal damage with or without metabolic activation.

Gene mutation (mammalian cell gene mutation assay):

There is one gene mutation (mammalian cell gene mutation assay) available.

In an in vitro gene mutation study in mammalian cells (Directive 87/302 EEC, L.133 p.61-63/GLP), CHO cells were exposed to 3,9-bis[2,4-bis(1-methyl-1-phenylethyl)phenoxy]-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane in DMSO at concentrations from 10 to 5000µg/mL for 4 hrs with Aroclor 1254-induced rat liver S9 metabolic activation and 10 to 3000µg/mL for 4 hours without metabolic activation. Cytotoxicity was observed at 5000 µg/ml with metabolic activation and 3000 µg/ml without metabolic activation in the preliminary tests. Cytotoxicity was observed at 5000 µg/ml with metabolic activation and ≥300 µg/ml without metabolic activation in the main tests. Precipitation was observed at ≥300 μg/ml with and without metabolic activation. There was no evidence of induced mutant colonies over background with or without metabolic activation.

All three tests are clearly negative and are suitable to use in the human health risk assessment.

Justification for classification or non-classification

Based on the available information, the substance the substance 3,9-bis[2,4-bis(1-methyl-1-phenylethyl)phenoxy]-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane (CAS No. 154862 -43-8) does not need to be classified for germ cell mutagenicity when the criteria outlined in Annex I of 1272/2008/EC) are applied.