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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
genetic toxicity in vitro
Remarks:
Type of genotoxicity: other: No-genotoxicity methods
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well-documented publication report which meets basic scientific principles.
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
publication
Title:
Investigation of cytotoxic,genotoxic,mutagenic,andestrogenic effects of the flame retardants tris-(2-chloroethyl)-phosphate (TCEP) and tris-(2-chloropropyl)-phosphate(TCPP) in vitro
Author:
W.Follmann, J.Wober
Year:
2005
Bibliographic source:
ToxicologyLetters161(2006)124–134

Materials and methods

Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
The NRU cytotoxicity assay procedure is a cell survival/viability chemosensitivity assay based on the ability of viable cells to incorporate and bind neutral red (NR), a supravital dye. NR is a weak cationic dye that readily penetrates cell membranes by non-ionic diffusion and accumulates intracellularly in lysosomes. Alterations of the cell surface or the sensitive lysosomal membrane lead to lysosomal fragility and other changes that gradually become irreversible. Such changes brought about by the action of xenobiotics result in a decreased uptake and binding of NR. It is thus possible to distinguish between viable, damaged, or dead cells, which is the basis of this assay.
Healthy mammalian cells, when maintained in culture, continuously divide and multiply over time. A toxic chemical, regardless of site or mechanism of action, will interfere with this process and result in a reduction of the growth rate as reflected by cell number. Cytotoxicity is expressed as a
concentration dependent reduction of the uptake of the NR after chemical exposure thus providing a sensitive, integrated signal of both cell integrity and growth inhibition.


GLP compliance:
not specified
Type of assay:
other: Cell Cytotoxicity study

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
tris-(2-chloropropyl)- phosphat(TCPP) (CASNo.:13674-84-5)was from Akzo-Nobel (Deventer, Netherlands).
There are differences in the isomer content from each supplier,but these are not important given that the properties of the isomers are expected to be very similar.
Purity
A typical purity(total of the four isomers)is>97.9%.All testing described in this report is for the commercial product.
Impurities
The impurity profile of the commercial product TCPP is specific to individual manufacturers. It is not likely that the impurities will have had particular influence on any of the results obtained.
Additives
No additives are used.

Method

Target gene:
Non-genotoxicity test
Species / strain
Species / strain:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell lines (if applicable):
In 96-well culture plates 15,000 cells per well were seeded in 200 μl culture medium. After 24 h culture mediumwas withdrawn, and the cells were incubated for 24 h with concentrations between 100pM and 10mM of the test substances in serum-free culture medium.
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9-rat liver homogenate
Test concentrations with justification for top dose:
100pM to 10mM TCPP
Vehicle:
DMSO
Controls
Negative controls:
no
Solvent controls:
yes
Remarks:
0.5% DMSO
True negative controls:
no
Positive controls:
no
Positive control substance:
no
Details on test system and conditions:
After 24 h culture mediumwas withdrawn, and the cells were incubated for 24 h with concentrations between 100pM and 10mM of
the test substances in serum-free culture medium. Stock solutions of the test substances were dissolved in DMSO. The final DMSO concentration in the incubation medium for all samples was 0.5%. As a solvent control cells were incubated with 0.5% DMSO. Routinely, a medium control
without any supplements and a growth control, from which the medium was withdrawn at the beginning of the incubation period, were performed in all tests.
Evaluation criteria:
no data
Statistics:
no data

Results and discussion

Test results
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
not determined
Cytotoxicity:
yes
Remarks:
above 1 mM with S9 and above 10mM without S9
Vehicle controls valid:
yes
Negative controls valid:
not specified
Positive controls valid:
not applicable
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
Nothing to add.

Any other information on results incl. tables

In the neutral red uptake assay TCPP showed a moderate toxicity in V79 cells at concentrations above 1 mM,in the presence of the external metabolizing enzyme system (S9-mix). In the absence of S9-mix no cytotoxic effect could be detected up to concentrations of 10mM. Parallel measurement of the protein content of the cells showed no impact on protein content under both conditions.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
other: positive results displayed above 1 mM with S9 and 10 mM without S9.

In the neutral red uptake assay TCPP showed a moderate toxicity in V79 cells at concentrations above 1 mM,in the presence of the external metabolizing enzyme system (S9-mix). In the absence of S9-mix no cytotoxic effect could be detected up to concentrations of 10mM. Parallel measurement of the protein content of the cells showed no impact on protein content under both conditions.