Bis(2-ethylhexyl) hydrogen phosphate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Bis(2-ethylhexyl) hydrogen phosphate was initially investigated using the Salmonella/microsome plate incorporation test for point mutagenic effects in doses of up to and including 5 µL per plate on five Salmonella typhimurium L T2 mutants. These comprised the histidine-auxotrophic strains TA 1535, TA 100, TA 1537, TA 98 and TA 102. The independent repeat was performed as preincubation for 60 minutes at 37°C. Other conditions remained unchanged.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Bis(2-ethylhexyl) hydrogen phosphate

EC Number:

206-056-4

EC Name:

Bis(2-ethylhexyl) hydrogen phosphate

Cas Number:

298-07-7

Molecular formula:

C16H35O4P

IUPAC Name:

bis(2-ethylhexyl) hydrogen phosphate

Test material form:

other: liquid

Details on test material:

Content: 99.3 %

Method

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Additional strain / cell type characteristics:

other: Firstly, they are deep rough since certain lipopolysaccharide side chains are missing in the bacterial cell wall. Secondly, their reduced ability to repair damage from UV light (e.g. thymidine dimers) allows the phenotypic detection of mutation events

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

Plate incorporation method: with and without S9 mix: 0, 0.05, 0.16, 0.5, 1.6, 5.0 µl/plate
Preincubation method: with and without S9 mix: 0, 0.05, 0.16, 0.5, 1.6, 5.0 µl/tube

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

2-nitrofluorene

sodium azide

cumene hydroperoxide

mitomycin C

other: anthracene-2-amine, 4-nitro-o-phenylenediamine

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

above 16 µl/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: other: S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

No evidence of mutagenic activity of bis(2-ethylhexyl) hydrogen phosphate was seen in this ames test. No biologically relevant increase in the mutant count, in comparison with the negative control, was observed up to the highest assessable doses.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

Executive summary:

Bis(2-ethylhexyl) hydrogen phosphate was initially investigated using the Salmonella/microsome plate incorporation test for point mutagenic effects in doses of up to and including 5 µL per plate on five Salmonella typhimurium L T2

mutants. These comprised the histidine-auxotrophic strains TA 1535, TA 100, TA 1537, TA 98 and TA 102. The independent repeat was performed as preincubation for 60 minutes at 37° C. Other conditions remained unchanged.

No precipitation was observed up to the highest tested dose.

In the plate incorporation test doses of up to and including 0.16 µl per plate did not cause any bacteriotoxic effects. At higher doses, bis(2-ethylhexyl) hydrogen phosphate had a strain-specific bacteriotoxic effect. Due to this effect this range could not be entirely used up to 5 µl per plate for assessment purposes.

In the preincubation test doses of up to and including 0.16 µl per plate did not cause any bacteriotoxic effects. At higher doses, bis(2-ethylhexyl) hydrogen phosphate had a strain-specific bacteriotoxic effect. Due to this effect this range could not be entirely used up to 5 µl per plate for assessment purposes.

Evidence of mutagenic activity of bis(2-ethylhexyl) hydrogen phosphate was not seen. No biologically relevant increase in the mutant count, in comparison with the negative control, was observed up to the highest assessable doses.

The positive controls sodium azide, anthracene-2-amine, cumene hydroperoxide, mitomycin C, 2-nitro-9H-fluorene and 4-nitro-o- phenylenediamine showed the expected mutagenic effect, as was seen by a biologically relevant increase in mutant colonies compared to the corresponding negative controls.

Therefore, bis(2-ethylhexyl) hydrogen phosphate was considered negative without and with S9 mix in the plate incorporation as well as in the preincubation modification of the Salmonella/microsome test.