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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.5300 - In vitro Mammalian Cell Gene Mutation Test
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
mammalian cell gene mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
- Name of test material (as cited in study report): Polyether V 531
- Molecular weight: 356
- Physical state: liquid
- Stability under test conditions: analytically approved
- content: 100%

Method

Target gene:
HPRT locus
Species / strain
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
Type and identity of media: PAA Ready Mix (2 % FBS)
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254 induced male rat liver S9 mix
Test concentrations with justification for top dose:
initial assay and independent repeat assay: 125, 250, 500, 1000, 2000, 3600 µg/ml (+ S9/-S9 mix)
Vehicle / solvent:
Vehicle(s)/solvent(s) used: water
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: ethylmethanesulfonate (EMS, 900 µg/ml, -S9); dimethylbenzanthracene (DMBA, 20 µg/ml, +S9)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 5 hours
- Expression time (cells in growth medium): 6-8 days

NUMBER OF REPLICATIONS: at least 2

DETERMINATION OF CYTOTOXICITY
- Method: cloning efficiency


Evaluation criteria:
Mutant frequencies will only be used for assessment, if at least 5 dishes per culture were available and relative survival to treatment, relative population growth and absolute cloning efficiency were 10% or greater.
- A trial will be considered positive if a concentration-related and in parallel cuJtures reproducible increase in mutant frequencies is observed. To be relevant, the increase in mutant frequencies should be at least two to three times that of the highest negative or negative control value observed in the respective trial. Ifthis result can be reproduced in a second trial, the test substance is considered to be mutagenic.
Despite these criteria, a positive result will only be considered relevant, if no significant change in osmolality compared to the negative control can be observed. Otherwise, unphysiological culture conditions may be the reason for the positive
result.
- A test substance will be judged as equivocal if there is no strictly concentration related increase in mutation frequencies but if one or more concentrations induce a reproducible and biologically relevant increase in mutant frequencies in all trials.
- An assay will be considered negative if no reproducible and relevant increases of mutant frequencies were observed.
However, these criteria may be overruled by good scienti'fic judgement.
Statistics:
All acceptable groups are included in the weighted analysis of variance followed by pairwise comparisons to the negative control on a nominal significance level of a =0.05 using the Dunnett test (Dunnett, C.W. (1955). J. Am. Sta1. Ass. 50,1096-1121). The regression analysis part is performed on the basis of the actual concentrations thereby omitting the positive, untreated and negative controls. If there is a significant concentration related increase of the mutant frequency (a = 0.05) in the main analysis the highest concentration will be dropped and the analysis will be repeated. This procedure will be repeated until p > 0.05.

Results and discussion

Test results
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

The test substance was evaluated for point mutagenic effects at the hypoxanthine-guanine phosphoribosyl transferase locus (forward mutation assay) in V79 cell cultures after treatment with concentrations of up to and including 3600 µg/ml, both with and without S9 mix.Without and with S9 mix the test substance induced no decreases in survival to treatment or in relative population growth. Precipitation of the test substance in the culture medium was not observed. However, 2,2',2''-Nitrilotriethanol, propoxylated was tested up to at least the limit concentration of 10 mM which is equal to 3560 µg/ml. Without and with S9 mix there was no biologically relevant increase in mutant frequency above that of the negative controls. Ethylmethanesulfonate and dimethylbenzanthracene induced clear mutagenic effects and demonstrated the sensitivity of the test system and the activity of the S9 mix. Based on these results, the test substance is considered to be non-mutagenic in the V79/HPRT forward mutation assay, both with and without metabolic activation.

Applicant's summary and conclusion

Conclusions:
non-mutagenic
Executive summary:

The test material was evaluated for point mutagenic effects at the hypoxanthine-guanine phosphoribosyl transferase locus (forward mutation assay) in V79 cell cultures after treatment with concentrations of up to and including 3600 µg/ml, both with and without S9 mix. The test substance induced no decreases in survival to treatment or in relative population growth in either the absence or presence of S9 mix when tested up to the limit concentration of 10 mM (equivalent to 3560 µg/mL. Precipitation of the test substance in the culture medium was not observed. There were no biologically relevant increases in mutant frequency in either the absence or presence of S9 mix above that of the negative controls. The positive controls induced clear mutagenic effects and demonstrated the sensitivity of the test system and the activity of the S9 mix.

Based on this study, the test material did not induce mutagenic effects in the in vitro gene mutation assay (OECD TG 476; HPRT test) with Chinese hamster V79 cells in the presence and absence of a metabolic activation system.