Tris(2-chloro-1-methylethyl) phosphate

Genetic toxicity: in vitro

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 for cross-reference:

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

Test material information

Constituent 1

Reference substance name:

Tris(2-chloro-1-methylethyl) phosphate

EC Number:

237-158-7

EC Name:

Tris(2-chloro-1-methylethyl) phosphate

Cas Number:

13674-84-5

Molecular formula:

C9H18Cl3O4P

IUPAC Name:

tris(2-chloro-1-methylethyl) phosphate

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 / cell type:

Chinese hamster lung fibroblasts (V79)

Details on mammalian cell type (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 / cell type 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 / solvent:

DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

0.5% DMSO

True negative controls:

no

Positive controls:

no

Positive control substance:

no

Details on test system and experimental 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 / choice of top concentrations:

cytotoxicity

Remarks:

above 1 mM with S9 and above 10mM without S9

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

not applicable

Additional information on results:

Nothing to add.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

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.