Reaction mass of ethyl dodecanoate and ethyl hexadecanoate and ethyl tetradecanoate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

The information is reliable and adequate for covering the endpoint.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

1

Reference substance name:

1,4-dioxacyclohexadecane-5,16-dione

EC Number:

259-423-6

EC Name:

1,4-dioxacyclohexadecane-5,16-dione

Cas Number:

54982-83-1

Molecular formula:

C14H24O4

IUPAC Name:

1,4-dioxacyclohexadecane-5,16-dione

Test material form:

liquid

Method

Target gene:

S. typhimurium TA1535 hisG46 rfa uvrB; S. typhimurium TA1537 hisC3076 rfa uvrB; S. typhimurium TA 1538 hisD3052 rfa uvrB; S. typhimurium TA98 hisD3052 rfa uvrB pKMl01; S. typhimurium TA100 hisG46 rfa uvrB pKM101; E. coli WP2 uvrA trp

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium, other: TA 1535, TA 1537, TA 1538, TA 98 and TA 100

Additional strain / cell type characteristics:

not applicable

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced rats (S9 mix)

Test concentrations with justification for top dose:

Exp 1: with and without S9-mix, 10 dose levels between 9.8 and 5000 µg/plate
Exp 2 with and without S9-mix
for TA1535, TA1537 and WP2 uvrA: 6 dose levels between 39.1 and 12500 µg/plate
for TA1538, TA98 and TA100: 8 dose levels between 39.1 and 12500 µg/plate

Vehicle / solvent:

dimethyl sulphoxide (DMSO)

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

and 9-Aminoacridine, 2-Nitrofluorene and 2-Aminoanthracene

Details on test system and experimental conditions:

An aliquot of 0.1 ml of a 10 hour bacterial culture and 0.5 ml S9 mix or 0.5 ml 0.1 M phosphate buffer (pH 7.4) were placed in glass bottles. An aliquot of 100 µl of the test solution was added, followed immediately by 2 ml of histidine/tryptophan deficient agar. The mixture was thoroughly shaken and overlaid onto previously prepared petri dishes containing 25 ml minimal agar. Each petri dish was individually labelled with a unique code corresponding to a sheet, identifying the dish's contents. Plates were also prepared without the addition of bacteria in order to assess the sterility of the test substance, 59 mix and phosphate buffer. All plates were incubated at 37°C for 3 days. After this period the appearance of the background bacterial lawn was examined and revertant colonies counted using a Seescan Automatic Colony Counter.

Evaluation criteria:

Please see below.

Statistics:

no

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 6

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

Toxicity was observed towards all the strains at the top three dose levels, down to 625 µg/plate for TA1538 and down to 312.5 µg/plate for TA98 and TA100. In the second mutation test toxicity was observed towards all the strains at the top dose level and down to 312.5 µg/plate for TA1538, TA98 and TA100.
No substantial increases in revertant colony numbers of any of the tester strains were observed following treatment at any dose level, in the presence or absence of S9 mix, in either mutation test.

Applicant's summary and conclusion

Conclusions:

It is concluded that the results obtained with the test substance in Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100, and in the Escherichia coli strain WP2 uvrA, in the absence and presence of the S9-mix, indicate that the test substance is not mutagenic under the conditions of this test.

Executive summary:

In a study, performed in accordance with GLP and OECD guideline 471 and 472, the test substance was examined for its possible mutagenic activity in the bacterial reverse mutation test using the Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98, TA 100 and the Escherichia coli strain WP2uvrA, in the absence and presence of a liver fraction of Aroclor 1254-induced rats for metabolic activation (S9-mix).The test substance was diluted in dimethylsulphoxide (DMSO). Two independent mutation tests were performed. A top dose level of 5000 µg/plate, based on a preliminary test, was chosen for the first mutation study. Other dose levels used were a series of 2-fold dilutions of the highest concentration. A total of ten dose levels were used in the first test to ensure that sufficient non-toxic concentrations were assessed. Toxicity was observed towards all the strains at the top three dose levels, down to 625 µg/plate for TA1538 and down to 312.5 µg/plate for TA98 and TA100. For the second mutation test a top dose level of 12500 µg/plate was chosen and the bottom two dose levels not included for TA1535, TA1537 and WP2 uvrA. Toxicity was observed towards all the strains at the top dose level and down to 312.5 µg/plate for TA1538, TA98 and TA100. No evidence of mutagenic activity was seen at any dose level in either mutation test. The concurrent positive control compounds demonstrated the sensitivity of the assay and the metabolising activity of the liver preparations. It is concluded that the test substance shows no evidence of mutagenic activity in this bacterial system.