Sucrose and glycerol, reaction products with C12-18, C18unsatd. fatty acids

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:

1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1983

Deviations:

no

Principles of method if other than guideline:

Performed as described by Maron DM and Ames BN (Revised methods for the Salmonella mutagenicity test. Mutation Res. 113. 173-215, 1983).
The study was performed equivalent to the OECD guideline 471 of 1983. Therefore, the additional strain S. typhimurium TA 102 or E. coli WP2 was not tested as required by the current guideline.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Sucrose and glycerol, reaction products with C12-18, C18unsatd. fatty acids

Molecular formula:

Representative, generic structures are given in "structural formula" wherein R = H or fatty acid residue and R' = sucrose residue, glycerol residue, H or alkali. Additional citric acid resp. its salt is present.

IUPAC Name:

Sucrose and glycerol, reaction products with C12-18, C18unsatd. fatty acids

Test material form:

not specified

Method

Target gene:

his

Species / strain

Species / strain / cell type:

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

Additional strain / cell type characteristics:

other: rfa, uvrB-

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

Dose finding preliminary toxicity test:
9.8, 19.5, 39.1, 78.1, 156.3, 312.5, 625.0, 1250, 2500, 5000 µg/plate

Main test
6.4, 32, 160, 800, 4000 µg per plate

Vehicle / solvent:

Methanol/H2O = 1 : 2

Controls

Controls 1

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

N-ethyl-N-nitro-N-nitrosoguanidine

other: 9-Aminoacridinehydrochloride monohydrate; 4-nitro-1,2-phenylene diamine

Remarks:

without S9 mix

Controls 2

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

with S9 mix

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

SELECTION AGENT (mutation assays): histidine deficient medium

NUMBER OF REPLICATIONS: 4

DETERMINATION OF CYTOTOXICITY
- Method: background growth / number of revertants

Evaluation criteria:

A reproducible and dose related increase of mutant counts for at least one strain is considered to be positive. For TA 100, TA 98 and TA 1535 a twofold increase of revertants compared to the negative controls should be reached, whereas for TA 1537 a threefold increase should be reached. Otherwise the results are estimated as being negative.

The following criteria were used for the acceptance of an assay:
- The negative controls have to be within the expected range as defined by literature data (Maron and Ames 1983).
- The positive controls have to show sufficient effects as defined by the laboratories' experience.
- The titer determination must yield a sufficient bacterial density in the suspension.

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

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
A bacteriotoxic effect was observed in doses ranging from 312.5 µg to 5000 µg per plate. Therefore the following doses were chosen for the main test: 6.4, 32, 160, 800 and 4000 µg/plate. The test substance was diluted in Methanol/H2O 1 : 2.

The evaluation of the individual dose groups with respect to the parameters relevant for assessment (dose effect and doubling), showed no biologically relevant variations from the respective negative controls for all strains.

In the positive controls, the mutant counts increased to more than twice the amount compared to the negative controls. This showed that the system is highly sensitive.

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

Based on the results of this study it is concluded that test substance is not mutagenic in the Salmonella typhimurium reverse mutation assay (test strains: TA 98, TA 100, TA 1535, TA 1537).

Executive summary:

In a reverse gene mutation assay in bacteria equivalent to OECD Guideline 471 (Bacterial Reverse Mutation), strains TA 1535, TA 1537, TA 98, T 100 of S. typhimurium were exposed to Sucroglyceride C12-18, C18unsatd. in Methanol/H₂O = 1 : 2, at concentrations of: 6.4, 32, 160, 800, 4000 µg per plate in the presence and absence of mammalian metabolic activation.

Sucroglyceride C12-18, C18unsatd. was tested up to cytotoxic concentrations. No evidence of a mutagenic activity of the test substance was found. Neither a dose related doubling nor a biologically relevant increase in the mutant count with and without S9 mix in comparison with the negative control (solvent) was observed. The positive controls induced the appropriate responses in the corresponding strains.

 

Based on the results of this study it is concluded that Sucroglyceride C12-18, C18unsatd. is not mutagenic in the Salmonella typhimurium reverse mutation assay.

 

This study is classified as acceptable. It satisfies the requirement for Test Guideline OECD 471 (1983) for in vitro mutagenicity (bacterial reverse gene mutation) data.

The negative result from the first test was confirmed by a second independent experiment. The results of the positive control substances confirm the specifity of the tester strains as well as the full activity of the metabolizing system.

The adopted OECD TG 471 (1997) requires at least 5 test strains and the use of E. coli WP2 strains or Salmonella typhimurium TA 102 to detect certain oxidizing mutagens, cross-linking agents and hydrazines. However, Sucroglyceride C12-18, C18unsatd.

is not a highly reactive agent and is therefore not expected to be a cross-linking agent, has no oxidizing properties and is no hydrazine. Thus, a GLP test according to former versions of the Test Guideline without E. coli WP2 strains or Salmonella typhimurium TA 102 is considered as sufficient to evaluate the mutagenic activity of Sucroglyceride C12-18, C18unsatd. in this bacterial test system.