cyclomaltodextrin glucanotransferase

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Cyclomaltodextrin glucotransferase was tested for genotoxicity using the Ames assay.

No dose-related and reproducible increases in revertants to prototrophy were obtained with any of the bacterial strains exposed to CGTase, either in the presence or absence of S-9 mix. It was concluded, that the results of the experiments, described in this report, gave no indication of mutagenic activity of CGTase, batch PPA 4357 in the presence or absence of metabolic activation.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10-01-1994 to 10-05-1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Target gene:

Histidine and tryptophan locus in the genome of four strains of bacteria.

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S-9 mix from Aroclor 1254-induced rat liver

Test concentrations with justification for top dose:

Preliminary test: No preliminary trials were carried out.
Experiment 1: 33, 100, 333, 1000, 3333, 10000 μg/mL.
Experiment 2: 313, 625, 1250, 2500, 5000, 10000 μg/mL.

Vehicle / solvent:

Vehicle for enzyme: Purified water.
Justification for choice of solvent/vehicle: Substance is water-soluble and any human exposure will be in aqueous solutions.
Solvent for the positive control: Sterile water.

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

other: N-methyl-N'-nitro-nitroso-guanidine, 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: Preincubation in suspension, followed by plating on agar plates (treat and plate method)
- Cell density at seeding (if applicable): Not stated

DURATION
- Exposure duration, pre-incubation: The incubation mixtures were incubated with shaking at 37 ± 1°C for 3 hours (treat and plate).
- Incubation time (selective incubation): 64 hours

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: Viable cell count and observation of the bacterial background lawn. 0.1 ml aliquots of a 10^6 dilution of each bacterial suspension were poured on to Nutrient agar plates.

Evaluation criteria:

A test substance is considered as positive in the Salmonella reverse mutation assay when it has induced a statistically significant increase in revertant count compared with the appropriate solvent control in one or more strains of bacteria, in the presence
or absence of S9, if this response is dose-related or reproduced in an independent experiments.

Statistics:

The numbers of revertants pr. plate were analysed by one-way analysis of variance for each test organism and each test series (1-way ANOVA performed 16 times). If the 1-way ANOVA gave some indications of significant intergroup differences, Dunnett's T-test were applied (one tail ed testing for any treatments significantly larger than the solvent control). The overall level of significance was alpha = 0.05.
All analysis were performed by means of the SAS statistical software package (PROC GLM).

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility: Yes
- Precipitation: No issues were reported in this study.
- Definition of acceptable cells for analysis: Viability and gene type control

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data: Yes
- Negative (solvent/vehicle) historical control data: Not stated

Some statistical significance in reverant colonies was attained for TA100 in the presence of S-9 mix for the 3 highest concentration in experiment 1, however, this was not reproducible in experiment 2.

Conclusions:

No dose-related and reproducible increases in revertants to prototrophy were obtained with any of the bacterial strains exposed to CGTase, either in the presence or absence of S-9 mix. It was concluded, that the results of the experiments, described in this report, give no indication of mutagenic activity of CGTase, batch PPA 4357 in the presence or absence of metabolic activation.

Executive summary:

CGTase, batch PPA 4357 was examined for mutagenic activity using Salmonella typhimurium strain TA1535, TA100, TA1537 and TA98. A liquid culture assay was applied. Bacteria were exposed to 6 doses of the test substance up to 10000 μg/mL in a phosphate buffered nutrient broth for 3 hours. After incubation, the test substance was removed by centrifugation prior to plating. The number of revertants to prototrophy and viable cells were estimated. The test was conducted in the presence and absence of metabolic activation - a liver preparation from male rats, pre-treated with Aroclor 1254, and the co-factors required for mixed function oxidase activity (S-9 mix). The sensitivity of the individual bacterial strains was confirmed by significant increases in number of revertant colonies induced in similar conditions by diagnostic mutagens. All results were confirmed by conducting two independent experiments.

No dose-related and reproducible increases in revertants to prototrophy were obtained with any of the bacterial strains exposed to CGTase, either in the presence or absence of S-9 mix. It was concluded, that the results of the experiments, described in this report, give no indication of mutagenic activity of CGTase, batch PPA 4357 in the presence or absence of metabolic activation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Justification for classification or non-classification

Not classified.