Glutens, hydrolyzates, reaction products with lauroyl chloride, sodium salts

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:

1998-04-29 to 1998-05-21

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study, OECD guideline

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

not specified

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Glutens, hydrolyzates, reaction products with lauroyl chloride, sodium salts

EC Number:

500-500-3

EC Name:

Glutens, hydrolyzates, reaction products with lauroyl chloride, sodium salts

Cas Number:

161074-67-5

Molecular formula:

Not applicable

IUPAC Name:

Acylation product between lauroyl chloride and amino acids

Test material form:

other: Liquid

Details on test material:

Sponsor’s identification : PROTÉOL OAT
Batch number : 98 014 007
Purity: 29.8% (dry extract)
Description : extremely paie yellow viscous liquid
Date received : 24 April 1998
Storage conditions : room temperature in the dark

Specific details on test material used for the study:

Batch Number: 98 014 007
Description: extremely pale yellow viscous liquid
Storage conditions: room temperature in the dark

Method

Target gene:

Histidine
Tryptophan

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Details on mammalian cell type (if applicable):

Salmonella strains were obtained from the Uiversity of California at Berkeley
Escherichia coli strain WP2uvrA- was obtained from the British Industrial Biological Research association
All of the strains were stored at -196°C

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

Preliminary study:
0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 pg/plate

Mutation Study
0 / 50 / 150 / 500 / 1500 / 5000 µg/plate

Vehicle / solvent:

Sterile distilled water

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

3 pg/plate for TA100, 5 pg/plate for TA1 535 and 2 pg/plate for E.coli WP2uvrA-

Controls 2

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

80 ug/plate for TA1537

Controls 3

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

0.2 pg/plate for TA98

Controls 4

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene

Remarks:

with S9-mix_ 1 µg/plate for TA100; 2 µg/plate for TA1535 and TA1537; 10µg/plate for WP2uvrA-

Controls 5

Positive control substance:

benzo(a)pyrene

Remarks:

with S9-mix_ 5 µg/plate for TA98

Details on test system and experimental conditions:

Preliminary toxicity study:
In order to select appropriate dose levels for use in the main study, a preliminary test was carried out to determine the toxicity of the test material. A mixture of 0.1 mL of bacterial culture (TA100 or E.coli), 2 mL of molten, trace histidine or tryptophan supplemented, top agar, 0,1 mL of test material formulation and 0.5 mL os S9-mix or phosphate buffer was overlaid onto sterile plates of Vogel-Bonner Minimal agar. Ten concentrations of the test material formulation and a vehicule control (streile distilled water) were tested. In addition, the sterility of the test material was assessed.
After approximately 48 hours incubation at 37°C the plates were assessed for numbres of revertatn colonies using a Domino colony counter.
Mutation Study-Experiment 1
Five concentrations of the test material were assayed in triplicate against each tester strain, using direct plate incorporation method.
All of the plates were incubated at 37°C for approximately 48 hours and the frequency of revertant colonies assessed using a Domino colony counter.
Mutation Study-Experiment 2:
The second experiment was performed using the same methodology as for the experiment 1.

Evaluation criteria:

The test material may be considered to be positive in this test system if the following criteria are met:
The test material should have induced a reproductible, dose related and Statistically significant increase in the revertant count in at least one strain of bacteria. If a greater than twofold increase in revertant count is observed in two experiments then this is taken as evidence of a positive response.

Statistics:

Dunnett's method of linear regression

Results and discussion

Test results

Species / strain:

other: all cell type tested

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

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

No significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains, with any dose of the test material, either with or without metabolic activation. The test material was considered to be non-mutagenic under the conditions of this test.

Executive summary:

Salmonella typhimurium strains TA1535, TA1537, TA98, TA100 and Escherichia coli strain WP2uvrA- were treated with the test material using the Ames plate incorporation method at five dose levels, in triplicate, both with and without the addition of a rat liver homogenate metabolising system (10% liver S9 in standard co-factors). This method conforms to the guidelines for bacterial mutagenicity testing published by the major Japanese Regulatory Authorities including MITI, MHW, MOL and MAFF. It also meets the requirements of the OECD, EC and USA, EPA (TSCA) guidelines. The dose range was determined in a preliminary toxicity assay and was 50 to 5000 pg/plate in the first experiment. The experiment was repeated on a separate day using the same dose range as experiment 1, fresh cultures of the bacterial strains and fresh test material formulations.

The vehicle (sterile distilled water) control plates gave counts of revertant colonies within the normal range.

All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies, both with and without metabolic activation. Thus, the sensitivity of the assay and the efficacy of the S9-mix were validated.

The test material caused a visible reduction in the growth of the bacterial lawn and/or a reduction in the frequency of revertant colonies to all of the Saimonella strains at the maximum recommended dose both with and without metabolic activation. The test material was, therefore, tested up to the maximum recommended dose of 5000 ug/plate.

No significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains, with any dose of the test material, either with or without metabolic activation. The test material was considered to be non-mutagenic under the conditions of this test.