Reaction product of "Fatty acids, C16-18 (even numbered) and C18-unsaturated" and "reaction mass of 1,3-alkanediol, 2-(hydroxymethyl)-2-[(methoxymethoxy)methyl]- and 1,3-heteromonocycle-5,5-dimethanol

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Link to relevant study records

Reference

Reference 1

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:

July 12-23, 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD guidelines and according to GLP principles

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Target gene:

- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene

Species / strain / cell type:

bacteria, other: Salmonella typhimurium (TA1535, TA1537, TA98 and TA100) and Escherichia coli (WP2uvrA)

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9 mix induced by phenobarbital/beta-naphthoflavon e

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 33, 100, 333, 1000 and 3330 µg/plate
Concentration range in the main test (without metabolic activation): 33, 100, 333, 1000 and 3330 µg/plate

Vehicle / solvent:

Solvent: Ethanol

Details on test system and experimental conditions:

Concentration of the test substance resulting in precipitation: 1000 µg/plate

Key result

Species / strain:

other: Salmonella typhimurium (TA100) and Escherichia coli (WP2uvrA)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

other: Salmonella typhimurium (TA1535, TA1537 and TA98)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

other: Salmonella typhimurium (TA1535, TA1537, TA98 and TA100) and Escherichia coli (WP2uvrA)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Observations:
In both mutation assays, there was no reduction of the
bacterial background lawn and no biologically relevant
decrease in the number of revertants at any of the
concentrations tested in all tester strains in the absence
and presence of S9-mix.

In both mutation assays, no increase in the number of
revertants was observed upon treatment with the test
substance under all conditions tested.

Remarks on result:

other: preliminary test

Comments:
The negative and strain-specific positive control values
were within our laboratory historical control data ranges
indicating that the test conditions were adequate and that
the metabolic activation system functioned properly.

Conclusions:

All bacterial strains showed negative responses over the entire dose range, i.e. no significant dose-related increase in the number of revertants in two independently repeated experiments.

Based on the results of this study it is concluded that Reaction products of Fatty acids, C16-18 and C18-unsatd. and reaction mass of 1,3-alkanediol, 2-(hydroxymethyl)-2-[(methoxymethoxy)methyl]- and 1,3-heteromonocycle-5,5-dimethanol is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

August 01 - October 14, 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD guidelines and according to GLP principles

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Species / strain / cell type:

mammalian cell line, other: Peripheral human lymphocytes

Metabolic activation:

with and without

Metabolic activation system:

phenobarbital and beta-naphthoflavone induced rat liver S9-m ix

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 10, 33 and 100 µg/ml
Concentration range in the main test (without metabolic activation): 10, 33 and 100 µg/ml
Concentration range in the main test (without metabolic activation): 10, 33, 100 and 333 µg/ml

Vehicle / solvent:

Ethanol

Details on test system and experimental conditions:

Exposure period (with metabolic activation): 3 hours
Exposure period (without metabolic activation): 3 hours

Fixation time:
Test 1:

24h (3h exposure) with and without S9


Test 2:

24h (24h exposure) and 48h (48h exposure) without S9

48h (3h exposure) with S9

Species / strain:

mammalian cell line, other: Peripheral human lymphocytes

Metabolic activation:

with and without

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Key result

Species / strain:

mammalian cell line, other: Peripheral human lymphocytes

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Observations:
Both in the absence and presence of S9-mix the test
substance did not induce a statistically significant or
biologically relevant increase in the number of cells with
chromosome aberrations in two independent experiments.

No effects on the number of polyploid cells and cells with
endoreduplicated chromosomes were observed both in the
absence and presence of S9-mix. Therefore it can be
concluded that the test substance does not disturb mitotic
processes and cell cycle progression and does not induce
numerical chromosome aberrations under the experimental
conditions described in this report.

Remarks on result:

other: preliminary test

Comments:
The number of cells with chromosome aberrations found in the
solvent control cultures was within the laboratory
historical control data range. The number of polyploid cells
and cells with endoreduplicated chromosomes in the solvent
control cultures was within the laboratory historical
control data range. The positive control chemicals (MMC-C
and CP) both produced statistically significant increases in
the frequency of aberrant cells. It was therefore concluded
that the test conditions were adequate and that the
metabolic activation system (S9-mix) functioned properly.

Conclusions:

It is concluded that Reaction products of Fatty acids, C16-18 and C18-unsatd. and reaction mass of 1,3-alkanediol, 2-(hydroxymethyl)-2-[(methoxymethoxy)methyl]- and 1,3-heteromonocycle-5,5-dimethanol is not clastogenic in human lymphocytes under the experimental conditions described in the report.

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29-Oct-2010 to 14-Dec-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

no

Principles of method if other than guideline:

The recommendations of the “International Workshop on Genotoxicity Tests Workgroup” (the IWGT), published in the literature (Clive et al., 1995, Moore et al., 1999, 2000, 2002, 2003, 2006 and 2007).

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian cell gene mutation assay

Target gene:

Thymidine kinase (TK) locus in L5178Y mouse lymphoma cells

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

- Type and identity of media:
- RPMI 1640 Hepes buffered medium (Dutch modification) containing penicillin/streptomycin (50 U/ml and 50 μg/ml, respectively), 1 mM sodium pyruvate and 2 mM L-glutamin supplemented with 10% (v/v) heat-inactivated horse serum (=R10 medium).
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: no
- Periodically "cleansed" against high spontaneous background: yes

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

Dose range finding test:
Without and with S9-mix, 3 hours treatment: 3, 10, 33, 100 and 500 µg/mL
Without S9-mix, 24 hours treatment: 3, 10, 33, 100 and 500 µg/ml
Experiment 1:
Without S9-mix, 3 hours treatment: 0.3, 1, 3, 10, 33, 100, 150 and 200 µg/mL
With S9-mix, 3 hours treatment: 0.1, 1, 3, 10, 33, 100, 300, 450 µg/mL
Experiment 2
Without S9-mix, 24 hours treatment: 0.3, 1, 3, 10, 33, 170, 200 and 220 µg/mL
With S9-mix, 3 hours treatment: 0.3, 1, 3, 10, 33, 100, 400 and 500 μg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Test compound was stable in DMSO and a suspension was obtained. DMSO has been accepted and approved by authorities and international guidelines

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

without S9; 15 µg/mL for the 3 hours treatment period and 5 µg/mL for the 24 hours treatment period

Positive control substance:

cyclophosphamide

Remarks:

with S9; 7.5 µg/mL

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration:
Short-term treatment
With and without S9-mix: 3 hours
Prolonged treatment period
Without S9-mix: 24 hours
- Expression time (cells in growth medium): 2 days
- Selection time (if incubation with a selection agent): 11 to 12 days

SELECTION AGENT (mutation assays): 5 µg/mL trifluorothymidine (TFT)

NUMBER OF REPLICATIONS:
- Solvent controls: Duplicate cultures
- Treatment groups and positive control: Single cultures

NUMBER OF CELLS EVALUATED: 9.6 x 10E5 cells plated/concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative suspension growth (dose range finding test) and relative total growth (mutation experiments)

Evaluation criteria:

Any increase of the mutation frequency should be evaluated for its biological relevance including a comparison of the results with the historical control data range. Furthermore:

The global evaluation factor (GEF) has been defined by the IWTG as the mean of the negative/solvent MF distribution plus one standard deviation. For the micro well version of the assay the GEF is 126.

A test substance is considered positive (mutagenic) in the mutation assay if it induces a MF of more than MF(controls) + 126 in a dose-dependent manner. An observed increase should be biologically relevant and will be compared with the historical control data range.

A test substance is considered equivocal (questionable) in the mutation assay if no clear conclusion for positive or negative result can be made after an additional confirmation study.

A test substance is considered negative (not mutagenic) in the mutation assay if:
a) None of the tested concentrations reaches a mutation frequency of MF(controls) + 126.
b) The results are confirmed in an independently repeated test.

Key result

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No
- Effects of osmolality: No
- Precipitation: Precipitation in the exposure medium was observed at dose levels of 100 µg/mL and above

RANGE-FINDING/SCREENING STUDIES:
- Toxicity was observed at the dose level of 500 µg/mL in the absence and presence of S9, 3 hours treatment; at the dose level of 500 µg/mL in the absence of S9, 24 hours treatment

COMPARISON WITH HISTORICAL CONTROL DATA:
The spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control data range and within the acceptability criteria of this assay.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
In the absence of S9-mix, the relative total growth of the highest test substance concentration was reduced by 81 and 85% compared to the total growth of the solvent controls after the 3 and 24 hours treatment period, respectively.



In the presence of S9-mix, the relative total growth of the highest test substance concentration was reduced by 85 and 72% compared to the total growth of the solvent controls after the 3 hours treatment period in the first and second experiment, respectively.

Remarks on result:

other: strain/cell type: L5178Y/TK+/-3.7.2C

Conclusions:

The spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control data range

Positive control chemicals, methyl methane sulfonate and cyclophosphamide induced appropriate responses.

In the absence of S9-mix, Reaction products of Fatty acids, C16-18 and C18-unsatd. and reaction mass of 1,3-alkanediol, 2-(hydroxymethyl)-2-[(methoxymethoxy)methyl]- and 1,3-heteromonocycle-5,5-dimethanol did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in a repeat experiment with modifications in the duration of treatment time.

In the presence of S9-mix, Reaction products of Fatty acids, C16-18 and C18-unsatd. and reaction mass of 1,3-alkanediol, 2-(hydroxymethyl)-2-[(methoxymethoxy)methyl]- and 1,3-heteromonocycle-5,5-dimethanol did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent experiment with modifications in the composition of the S9 concentration for metabolic activation.

In conclusion, Reaction products of Fatty acids, C16-18 and C18-unsatd. and reaction mass of 1,3-alkanediol, 2-(hydroxymethyl)-2-[(methoxymethoxy)methyl]- and 1,3-heteromonocycle-5,5-dimethanol is not mutagenic in the TK mutation test system under the experimental conditions described in the report.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Additional information

Ames test: In a study performed according to OECD and EC test guidelines, the species S. typhimurium TA1535, 1537, 98 and 100 and the species E. coli WP2 uvr A were tested up to the precipitating concentration of 3330 microg/plate with and without metabolic activation. All bacterial strains showed negative responses over the entire dose range with and without metabolic activation, i.e. no significant dose-related increase in the number of revertants in two independently repeated experiments. Based on the results of this study it is concluded that radia 7853 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Chromosome aberration test: In a study performed according to OECD and EC test guidelines, human lymphocytes were exposed to the test substance up to 100 µg/ml with and up to 333 µg/mL without metabolic activation, concentrations at which the substance precipitated. Radia 7853

did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of the two independently repeated experiments. Therefore it was concluded that the substance is not clastogenic in human lymphocytes under the experimental conditions chosen.

Mouse lymphoma test: In a study performed according to OECD and EC test guidelines, L5178Y mouse lymphoma cells were exposed to the test substance up to 220 µg/mL without metabolic activation and up to 500 µg/mL with metabolic activation. Precipitation was observed at 100 µg/mL

and above with and without metabolic activation. Toxicity was observed at the dose level of 500 µg/mL in the absence and presence of S9, 3 hours treatment; and at the dose level of 500 µg/mL in the absence of S9, 24 hours treatment. In the absence or presence of S9 -mix, the substance did not induce a significant increase in the mutation frequency. Based on the results of the test it was concluded that Radia 7853 is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions chosen.

Short description of key information:

The Ames study was performed according to OECD Guideline 471 and GLP principles. The chromosome aberration study was performed according to OECD Guideline 473 and GLP principles. The TK mutation test was performed according to OECD Guideline 476 and GLP principles.The substance tested 'negative' in all three tests performed.  

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the results of the genotoxicity studies, the substance does not need to be classified according to the CLP Regulation (EC) 1272/2008.