2,2,2,4,4,4,4-heptakis[(2-ethylhexanoyl)oxy]cyclodimolybdoxan-2-yl 2-ethylhexanoate

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

Study period:

16 February 2011 to 10 June 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was conducted in accordance with International Guidelines and in accordance with the OECD Principles of Good Laboratory Practice as incorporated into the United Kingdom (Statutory Instrument for GLP).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2,2,2,4,4,4,4-heptakis[(2-ethylhexanoyl)oxy]cyclodimolybdoxan-2-yl 2-ethylhexanoate

EC Number:

700-831-5

Molecular formula:

Mo2O2L8 (L = 2-ethylhexanoic acid; C8H16O2) (L' = 2-ethylhexanoic acid; C8H16O2)

IUPAC Name:

2,2,2,4,4,4,4-heptakis[(2-ethylhexanoyl)oxy]cyclodimolybdoxan-2-yl 2-ethylhexanoate

Method

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

S9 Mix

Test concentrations with justification for top dose:

In the absence of S9 mix, the concentrations tested were: 5, 17, 50, 167, 500 and 1667 μg per plate
In the presence of S9 mix, the concentrations tested were: 17, 50, 167, 500, 1667 and 5000 μg per plate

Vehicle / solvent:

Ethanol was used as the vehicle control and was plated in triplicate (50 μL per plate) with each strain used, both in the absence and in the presence of S9 mix.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: With S9 mix: 2-Aminoanthracene (2AAN): 2 μg per plate with S typhimurium TA 1535 and TA 1537, 0.5 μg per plate with S. typhimurium TA 98 and TA 100 and 20 μg per plate with E. coli WP2uvrA

Controls 2

Positive control substance:

other: Without S9 mix: Sodium azide (NaN3): 1 μg per plate with S. typhimurium TA 1535 and TA 100 9-Aminoacridine (9-AA): 80 μg per plate with S. typhimurium TA 1537

Controls 3

Positive control substance:

other: Without S9 Mix: 2-Nitrofluorene (2-NF): 1 μg per plate with S. typhimurium TA 98 N-Ethyl-N-nitro-N-nitrosoguanidine (ENNG): 2 μg per plate with E. coli WP2uvrA

Results and discussion

Test results

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

only in the absence of S9 mix

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

Toxicity Tests
In the absence of S9 mix, concentrations of 500 μg per plate and above were toxic to the bacteria. Toxicity was observed as a reduction in the number of revertant colonies and as a slight thinning of the background lawn of microcolonies. No precipitation of the test item occurred.
In the presence of S9 mix, no toxicity was observed. HCAT precipitated at the highest concentration of 5000 μg per plate.

Evidence of mutagenic activity was not obtained with HCAT in any strain in either test.
In the first mutation assay, conducted by the direct plate incorporation method, toxicity to the bacteria was observed in the absence of S9 mix as a slight thinning of the background lawn of microcolonies at the 2 highest concentrations of 500 and 1667 μg per plate. In the presence of S9 mix, toxicity to the bacteria was observed only at 5000 μg per plate in strain TA 1537. HCAT precipitated at 5000 μg per plate in the presence of S9 mix.
In the second mutation assay, conducted by the pre-incubation method, toxicity to the bacteria was observed in the absence of S9 mix at 1667 μg per plate in all strains and at 500 μg per plate in strains TA1535 and TA 1537. In the presence of S9 mix, toxicity to the bacteria was observed at 5000 μg per plate in all S. typhimurium strains. HCAT precipitated at 5000 μg per plate in the presence of S9 mix.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

In conclusion, HCAT was not mutagenic in strains of S. typhimurium and E. coli when dissolved and diluted in ethanol and tested in the absence and presence of metabolic activation. HCAT was tested at concentrations that extended into the toxic range in the absence of metabolic activation and up to the predetermined maximum concentration of 5000 μg per plate in the presence of metabolic activation. This latter treatment was beyond the limit of HCAT’s solubility in the test system.

Executive summary:

HCAT was tested for mutagenic activity in Salmonella typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and in Escherichia coli WP2uvrA.

Two independent tests were conducted on agar plates in triplicate in the absence and presence of an Aroclor 1254-induced rat liver S9 preparation and the co-factors required for mixed- function oxidase activity (S9 mix). The first test was conducted by the direct plate incorporation method, while the second test was conducted by the pre-incubation method. HCAT was dosed at concentrations ranging from 5 to 1667 μg per plate in the absence of S9 mix and from 17 to 5000 μg per plate in the presence of S9 mix. HCAT was dissolved and diluted in ethanol.

Concurrent positive controls demonstrated the sensitivity of the assay and the metabolising activity of the S9 mix.

Evidence of mutagenic activity was not obtained with any strain in either test.

Dependent on strain and test conditions, HCAT was toxic to the bacteria at the higher concentrations. This toxicity was observed either as a reduction in the number of revertant colonies, or as a thinning of the background lawn of microcolonies. Precipitation of HCAT occurred in the presence of S9 mix only, at the highest concentration of 5000 μg per plate.

In conclusion, HCAT was not mutagenic in strains of S. typhimurium and E. coli when dissolved and diluted in ethanol and tested in the absence and presence of metabolic activation. HCAT was tested at concentrations that extended into the toxic range in the absence of metabolic activation and up to the predetermined maximum concentration of 5000 μg per plate in the presence of metabolic activation. This latter treatment was beyond the limit of HCAT’s solubility in the test system.

The study was performed in accordance with the principles of Good Laboratory Practice.