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Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames test: Melamine phosphate was negative.


Chromosome Aberration assay: Melamine was negative.


Mammalian cell mutation assay: Melamine was negative.

Link to relevant study records

Referenceopen allclose all

Endpoint:
genetic toxicity in vitro, other
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
For Read Across Justification please refer to section 13.
Reason / purpose for cross-reference:
read-across source
Reason / purpose for cross-reference:
read-across source
Key result
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: No toxicity was observed, although some cell cycle delay was found in the SCE test.
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: CAS 108-78-1
Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2002
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
Purity >98%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature in the dark
- Solubility and stability of the test substance in the solvent/vehicle: at least 96 h in vehicle ethanol
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
liver S9 from Aroclor 1254-induced rats
Test concentrations with justification for top dose:
3,10, 33,100, 333, 1000, 3330 and 5000 µg/plate
Vehicle / solvent:
Ethanol
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene
Remarks:
for all strains with S9 mix
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
strain TA 1535 without S9 mix
Positive control substance:
9-aminoacridine
Remarks:
TA 1537 without S9 mix
Positive control substance:
other: daunomycine
Remarks:
TA 98 without S9 mix
Positive control substance:
methylmethanesulfonate
Remarks:
TA 100 without S9 mix
Positive control substance:
other: 4-nitroquinoline-N-oxide
Remarks:
WP2uvra without S9 mix
Details on test system and experimental conditions:
Based on the results of the dose range finding test, the test item was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix in two mutation assays. The first mutation experiment was performed with the strains TA 1535, TA 1537 and TA98 and the second mutation experiment was performed with the strains TA1535, TA1537, TA98, TA100 and WP2uvrA.
The pre-incubation assay was used.

Before use, all S9-batches were characterized with the metabolic activation requiring positive control; benzo[a]pyrene (Sigma) in tester strain TA98 at the concentration of 5 µg/plate.
Evaluation criteria:
A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in any tester strain at any concentration is not greater than two times the solvent control value, with or without metabolic activation.
b) The negative response should be reproducible in at least one independently repeated experiment.
A test substance is considered positive (mutagenic) in the test if:
a) It induces a number of revertant colonies, dose related, greater than two-times the number of revertants induced by the solvent control in any of the tester strains, either with or without metabolic activation. However, any mean plate count of less than 20 is considered to be not significant.
b) The positive response should be reproducible in at least one independently repeated experiment.
Key result
Species / strain:
other: TA 1535, TA1537, TA 98, TA 100
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:
not examined
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
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:
not examined
Positive controls validity:
valid
Additional information on results:
The test item precipitated in the top agar at all concentrations tested. Precipitation on the plates was observed at the start and at the end of the incubation period in all tester strains at the highest concentration tested.

The bacterial background lawn was not reduced at all concentrations tested and no biologically relevant decrease in the number of revertants was observed.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Micronucleus assay: Melamine was negative.

Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
For Read Across Justification please refer to section 13.
Reason / purpose for cross-reference:
read-across source
Reason / purpose for cross-reference:
read-across: supporting information
Key result
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Negative controls validity:
not applicable
Positive controls validity:
valid
Remarks on result:
other: CAS 108-78-1
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

In vitro

Melaminephosphate was tested in the Salmonella typhimurium reverse mutation assay according OECD TG 471 with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in the Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli WPrUvrA. The test was performed in two independent experiments in the presence and absence of S9-mix (Aroclor-1254 induced rat liver S9-mix). The substance was tested up to concentrations of 5 mg/plate in the absence and presence of S9-mix. It precipitated on the plates at this dose-level. The bacterial background lawn was not reduced at all concentrations tested and no biologically relevant decrease in the number of revertants was observed (BASF SE 335194, 2002).

A chromosome aberration study with melamine was performed according to the US NTP standard protocol which is similar to the procedure outlined in OECD testing guideline 473. Chinese hamster Ovary (CHO) cells were used together with and without S9 mix prepared from the liver of Aroclor-1254 induced rats. Applied doses were 240, 270 and 300 µg per plate. Mitomycin C and cyclophosphamide were used as positive controls. Melamine was found to be non clastogenic in vitro. Considering the higher molecular weight of melamine phosphate and that both are of comparable solubility in water, this data is adequate to conclude that melamine phosphate is not clastogenic in vitro (Galloway, 1987).

Mutagenicity in mammalian cells in vitro was assessed in the mouse lymphoma assay performed according the US National Toxicology Standard Program which is comparable to OECD testing guideline 476. Concentrations of melamine were 0; 10; 20; 40; 80; 100; 120; 140; 160 µg/mL ; testing of melamine was limited to 160 µg/mL by its solubility in the exposure medium. A dose-dependent increase in mutant frequency was observed for the positive control, but not for the solvent control DMSO and not for melamine. The cloning efficiency was not affected by melamine.Considering the higher molecular weight of melamine phosphate and that both are of comparable solubility in water, this data is adequate to conclude that melamine phosphate is not mutagenic to mammalian cells in vitro (McGregor 1988).

In vivo

Melamine was investigated in a micronucleus study in vivo. Groups of five mice were administered the substance at 0, 500, 1000 or 2000 mg/kg by intraperitoneal injection on three consecutive days. Two days after the third treatment, animals were sacrificed and bone marrow and peripheral blood smears prepared. Per animal, two slides per tissue were prepared. The initial test gave a positive trend (P = 0 .003) from 2.1 in the control to 3.8 in the high dose (2000 mg/kg). Peripheral blood smears from the dose determination test were scored and were negative as was a repeat bone marrow test using doses of 1000 and 2000 mg/kg (Shelby 1993).

In another micronucleus test according to Standard Operating Procedure PH-309A, melamine was administered orally to mice in a dose of 1000 mg/kg bw. Distilled water was used as vehicle control and triethylenemelamine as positive control. Erythrocytes were harvested from femurs after 30 h, 48h or 72 h and analyzed for micronuclei. The results for melamine were negative in the Micronucleus Test, at a dose of 1000 mg/kg in the single dose groups and at 2 x 1000 mg/kg administered in a split dose regimen to the multiple dose groups. This is based upon the inability of the chemical to produce a statistically significant increase in the number of micronuclei per 1000 polychromatic erythrocytes in the treated versus the control group (American Cynamide PH 309A-AC-001-81, 1981).

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008:


The available experimental test data are reliable and suitable for classification purposes. As a result the substance is not considered to be classified for genotoxicity under Regulation (EC) No. 1272/2008, as amended for the fourteenth time in Regulation (EU) No. 2020/217.