acetalization product between glucose and C16-18(even numbered)- alcohol

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:

1991/07/18 to 1991/10/28

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

guideline from 1983

Deviations:

yes

Remarks:

E coli or S. typhimurium TA 102 not performed. It was replaced by the strain TA1538

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Reference

Name:

Unnamed

Type:

Constituent

Method

Target gene:

Mutation in Histidine biosynthesis : his D 3052 (frameshift), his G 46 (base-pair substitution) and his C 3076 (frameshift)

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

0.1 ; 0.5 ; 1 ; 2.5 ; 5 mg/well

Vehicle / solvent:

absolute ethanol

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

for TA98 an TA1535 without S9 activation

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

for TA98, TA100, TA1535, TA1537, TA1538 with S9 activation

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

for TA100 without S9 activation

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

for TA1537 without S9

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

for TA21535 without S9

Details on test system and experimental conditions:

(1) Preliminary evaluation of cytotoxicity of the substance
The cytotoxicity was performed of the Salmonella Typhimurium strain TA 100 without metabolic activation. The substance was tested at 5 concentrations : 0.05 ; 0.1 ; 0.5 ; 1 ; 2.5 and 5 mg/dish. The incubation system contained 50 µl of the diluted substance + 2 ml of agar-upper layer + 0.1 ml of bacterial culture. A slight precipitate of the substance was observed at the concentration of 0.5 ; 1 ; 2.5 and 5 mg/dish. The mixture was plated onto a Petri dish.
Each concentration was tested twice. The Petri dishes were incubated at 37°C for 48 to 72 hours.
(2) Main study
5 strains of Salmonella Typhimurium were tested: TA98, TA100, TA1535, TA1537 and TA1538.
As the concentration of 5 mg/dish was not toxic in the strain TA 100 without metabolic activation during the preliminary study, the following concentrations were tested during the main study: 0.1 ; 0.5 ; 1 ; 2.5 and 5 mg/dish.
Each concentration was tested 3 times under a constant volume of 500 µl with and without metabolic activation (S9 fraction obtained from rat livers pre-treated with Aroclor 1254 at 500 mg/kg) and used at on each strain. The incubation system contained 50 µl of the diluted substance + 2 ml of agar-upper layer + 0.1 ml of bacterial culture +/- 0.5 ml of S9 mixture. The mixture was plated on a Petri dish and incubated at 37°C for 48 to 72 hours.
Absolute ethanol and positive controls, 2-nitrofluorene at 0.001 mg/dish for TA-98 and TA-1538, 2 aminoanthracene at 0.002 mg/dish for the TA-98, TA-100, TA-1535, TA-1537 and TA-1538 with S9 mixture, methyl methanesulfonate at 0.1 mg/dish for TA-100, ethyl methanesulfonate at 10 mg/dish for TA-1535, 9-aminoacridine at 0.05 mg/dish for TA-1537
All the results were confirmed in a second study independent from the first.

Evaluation criteria:

(1) Preliminary cytotoxicity
After incubation, the colonies were counted and the intensity of the bacterial lawn examined and compared with those performed with the vehicle alone. The signs of toxicity were noted.
(2) Reverse mutation
At the end of the incubation period, the colonies or revertants apparent in each dish were counted manually or automatically by an image analyser directly connected to a computer. Each count was then recorded and analysed directly.

Statistics:

calculation of the mean and standard deviation of n=3 per study.

Results and discussion

Test results

Species / strain:

S. typhimurium, other: TA98, TA100, TA1535, TA1537, TA1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

under experimental conditions employed, no value obtained in the presence of the test article was greater than or equal to twice the value obtained in the presence of the vehicle with and without metabolic activation on the bacterial strains used.
All the results obtained in presence of positive controls were significant with and without metabolic activation in the used bacterial strains.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'. Remarks: mutants of Salmonella typhimurium LT2

Applicant's summary and conclusion

Conclusions:

The test article is not mutagenic as it induces no significant increase in the number of revertants with and without metabolic activation of Salmonella Typhimurium TA98, TA100, TA1535, TA1537 and TA1538.

Executive summary:

The substance was tested on 5 strains of salmonella typhimurium (TA98, TA100, TA1535, TA1537 and

TA1538), with or without metabolic activation. A range of sub-toxic concentrations was determined in a preliminary study on the strain TA-100 without metabolic activation.

The 5 concentrations (0.1 - 0.5 - 1 - 2.5 - 5 mg/dish) were tested 3 times on the 5 strains mentioned above with and without metabolic activation. The results were confirmed in a second study, independant of the first.

In each study was included a negative control (vehicle = absolute ethanol) and a positive control (specific standard mutagen).

Under the experimental conditions employed, the substance did not show any mutagenic potential for the strain TA98, TA100, TA1535, TA1537 and TA1538 with and without metabolic activation.