1,3,5-tris(oxiranylmethyl)-1,3,5-triazine-2,4,6(1H,3H,5H)-trione

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:

1987

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: not stricty according to OECD 471, no statistical analysis

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1987

Report date:

1987

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

not stricty according to OECD 471, no statistical analysis

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

1,3,5-tris(oxiranylmethyl)-1,3,5-triazine-2,4,6(1H,3H,5H)-trione

EC Number:

219-514-3

EC Name:

1,3,5-tris(oxiranylmethyl)-1,3,5-triazine-2,4,6(1H,3H,5H)-trione

Cas Number:

2451-62-9

Molecular formula:

C12H15N3O6

IUPAC Name:

tris[(oxiran-2-yl)methyl]-1,3,5-triazinane-2,4,6-trione

Details on test material:

Araldite PT810 (TGIC), 1,3,5-triglycidylisocyanurate.
Purity: Commercial grade (> 97%)
Impurity < 100 ppm epichlorohydrin (CAS-no. 106-89-8)

Method

Target gene:

histidine auxotrophe

Species / strain

Species / strain 1

Species / strain / cell type:

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

Additional strain / cell type characteristics:

not applicable

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

S9- rat liver extract Arochlor 1254 induced

Test concentrations with justification for top dose:

1.22 - 10000 microgram/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

Migrated to IUCLID6: 2-nitrofluorene, quinacrine mustard, 2-anthramine

Details on test system and experimental conditions:

Positive control substances were Sodium azide (in water) for nonactivated TA-1535 & TA-100; 2-nitrofluorene (in DMSO) for nonactivated TA-1538 & TA-98; Quinacrine mustard (in DMSO) for nonactivated TA-1537; 2-Anthramine (in DMSO) for all activated strains
All strains are histine auxotrophs; only revertant mutations to histidine independence are growing in medium with minimal amounts of histidine. Spontaneous reverse mutations are rare, and only mutation rates of 2-100 fold are considered positive. Strains were received from Bruce Ames.
Media used were: Oxoid Media #2 (nutrient broth) for overnight growth at 37°C; Selection media for mutagenicity assay was Vogel Bonner Medium E (Vogen and Bonner, 1956) with 2% glucose and 1.5% bactoagar.; overlay medium was 0.6% purified agar, 10% of 0.5 mM L-Histidine and 0.5 mM Biotin, and 0.5% NaCl according to Ames et al (1975).
Assay were performed with and without S9 activating homogenate.
S9 was 9000 xg supernatant from Sprague-Dawley adult male rat liver induced by Aroclor 1254. The S9 mix consited of NADP (sadium salt) 4 μmoles, D-Glucose-6-phosphate 5 μmoles, MgCl2 8 μmoles, KCl 33 μmoles, Na-Phosphate buffer pH 7.4 100 μmoles, and S9 homogenate 100 μlitres
Dose selection was performed with TA-100 in the non-activated system in overlay agar contailing TGIC (0.05 -0.1 ml of appropriate solution), 0.1-0.2 ml indicator organisms, and 0.5 ml 0.2M phosphate buffer pH 7.4. This solution (at 43-45°C) was added to minimal agar plates and incubated at 37°C for two days. Colonies were counted manually on the plates. Reduction of colony count was considered as toxicity.
Main assay conduct:
Non-activated: 2 ml Overlay agar, 0.05-0.1 ml TGIC solution , 0.2 ml indicator organism, and 0.5 ml 0.2M Phosphate buffer pH 7.4, was swirled gently and poured over a minimal agar plate and incubated for 48 hours. Colonies were counted manually on the plates.
Activated: Same as non-activated , but instead of 0.5 ml =.2M Phosphate buffer pH 7.4, 0.5 ml of S9 mix is added.
Negative controls were run in parallel (only overlay agar with test organisms).
Strains TA-98 and TA-100: at least three concentrations in a row have to be positive, and the response has to be at least 2-times the control value.
Control values were : TA-1535: 8-30; TA-1537: 4-30; TA-1538: 10-40; TA-98: 20-75; TA-100: 80-250.
Experiments were performed in triplicates, and 2 independent experiments were performed.

Evaluation criteria:

Evaluation of data: Strains TA-1535, TA-1537, TA-1538: at least three concentrations in a row have to be positive, and the response has to be at least 3-times the control value.

Statistics:

No statistical analysis of the data was performed.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

ambiguous

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

The assay was positive (mutagenic activity associated with TGIC, araldite PT 810).

Doses assayed were from 1.22 to 10’000 μg/plate, showing some toxicity 5000 and complete toxicity at 10’000 μg/plate with TA-100 in the preliminary assay.

Therefore, doses of 1-10’000 μg/plate were used throughout the experiment with all strains.

The positive controls were within the expected range of induced colonies

Applicant's summary and conclusion

Conclusions:

TGIC caused mutagenic effects in all strains tested with the exception of TA 1538 (ambiguous results) in the Ames test, and thus, it is mutagenic in-vitro

Executive summary:

Araldite PT 810 (TGIC) was mutagenic in three different Salmonella strains (TA-1535, TA-98 and TA-100) in the presence and absence of S9-activating enzymes of rat liver induced with Aroclor 1254.Tehrefore, Araldite PT 810 is mutagenic in bacteria in-vitro. Based on the results of the non-activating system, it is obvious that TGIC is  a Direct-Acting Mutagen