4,4'-Methylenediphenyl diisocyanate, oligomeric reaction products with 2,4'-diisocyanatodiphenylmethane

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07.01.2013 - 04.02.2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Hypothesis: Under physiological conditions the MDI substance readily polymerizes at the MDI/aqueous interface forming insoluble polyureas and/or reacts with extracellular biological nucleophiles to form MDI-adducts rendering the free NCO completely unavailable to react with DNA thereby negating DNA reactivity concerns. Further, the toxicokinetic and metabolic pathways do not indicate the formation of toxicologically relevant metabolites.

Justification: Toxicokinetic and hydrolysis mechanisms demonstrate that MDI rapidly polymerizes to polyurea that the MDI/aqueous interface. Hydrolysis is highly unfavored except when reaction is stabilized by aprotic solvents (DMSO). When appropriate solvents are used, all tested MDI substances are negative for mutagenicity and is supported by this mechanism.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

PIR modified MDI

Cas Number:

109331-54-6

IUPAC Name:

PIR modified MDI

Method

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Metabolic activation system:

Type and composition of metabolic activation system:
- source of S9: liver S 9 fraction of Aroclor 1254-induced male Sprague Dawley rats
- concentration or volume of S9 mix and S9 in the final culture medium : plate-incorporation: 0.1 ml, pre-incubation 0.3 ml,
- quality controls of S9 (e.g., enzymatic activity, sterility, metabolic capability): yes

Test concentrations with justification for top dose:

0.01, 0.025, 0.05, 0.16, 0.5, 1.6, 5.0 µL/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: EGDE (dried with molecular sieve, 0.3 nm)
- Justification for choice of solvent/vehicle: test substance not stable in water or DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

sodium azide

mitomycin C

Remarks:

additional positive controls: anthracene-2-amine, 4-NPDA

Details on test system and experimental conditions:

NUMBER OF REPLICATIONS:
- Number of cultures per concentration: triplicate
- Number of independent experiments: two

METHOD OF TREATMENT/ EXPOSURE:
- Test substance added in medium; in agar plate incorporation and preincubation

Evaluation criteria:

The following criteria determined the acceptance of an assay:
1. The negative controls had to be within the expected range, as defined by published data
(e.g. Maron and Ames, Revised methods for the Salmonella mutagenicity test
Mutation Res. 1983, 113:173-215) and/ or the laboratories' own historical data
2. The positive controls had to show sufficient effects, as defined by the laboratories'
Experience
3. Titer determinations had to demonstrate sufficient bacterial density in the suspension.

Statistics:

mean, standard deviation

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

The test substance was considered to be non-mutagenic without and with S9 mix in the
plate incorporation as well as in the preincubation modification of the Salmonella/microsome
test.

Executive summary:

The mutagenic potential of the test substance was evaluated in a Salmonella/microsome test with the S. typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 in the presence and absence of S9 mix according to OECD TG 471. Evidence of mutagenic activity was not seen. No biologically relevant increase in the mutant count, in comparison with the negative controls, was observed. Based on this test, the test substance was considered to be non-mutagenic without and with S9 mix in the plate incorporation as well as in the preincubation modification of the Salmonella/microsome test.