m-tolyl isocyanate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

K2, AMES TEST, in vitro, Gahlmann (1994), Negative.

QSAR, AMES Test, Aromatic Amines, BFS (2022) Negative.

QSAR, AMES Test, Isocyanates, BFS (2022) Negative.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

(Q)SAR

Adequacy of study:

other information

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with limited documentation / justification

Justification for type of information:

AROMATIC AMINES

1. SOFTWARE
OECD QSAR Toolbox

2. MODEL (incl. version number)
v4.5

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
Cc1cccc(c1)N=C=O

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
[Explain how the model fulfils the OECD principles for (Q)SAR model validation. Consider attaching the QMRF and/or QPRF or providing a link]
see TPRF information below

5. APPLICABILITY DOMAIN
[Explain how the substance falls within the applicability domain of the model]
see TPRF information below

6. ADEQUACY OF THE RESULT
see TPRF information below

Prediction summary
Predicted endpoint: Gene mutation; No effect specified; No species specified; No duration specified; No guideline specified
Predicted value: Negative
Unit/scale: Gene mutation I
Data gap filling method: Read-across analysis
Summary:
The Ames mutagenicity of m-tolylisocyanate was predicted for the strains S. thyphimurium TA 102 or E. Coli WP2 UvrA, which were not present in existing Ames studies on the query compound. The Toolbox endpoint was specifically defined as being Genetic Toxicity specific to the strains S. thyphimurium TA 102 or E. Coli WP2 UvrA, To derive the prediction, a combination of supporting general mechanistic and endpoint specific profilers was used to assess the shared identified isocyanate acylation mechanism related to the query compound.
Similar compounds sharing these alerts were identified, and using the nearest neighbours to the query compound, a prediction of negative for Ames mutagenicity was derived, using only data from S. thyphimurium TA 102 or E. Coli WP2 UvrA, lending weight to the already establish experimental result on the query compound in other strains.

Predicted value: Negative
Predicted endpoint (OECD Principle 1 - Defined endpoint): Human Health Hazards -> Genetic Toxicity -> Bacterial Reverse Mutation Assay (e.g. Ames Test) -> Gene mutation -> E. Coli WP2 UvrA TA 102 WP2 UvrA
Prediction plot: (see Figure 1 below)

Calculation approach (OECD principle 2 - Unambiguous algorithm):
takes the highest mode value from the 4 nearest neighbours
Active descriptor: log Kow (calculated)
Data usage: Maximal value* *When multiple values are available for the same chemical, their maximal value is taken in prediction calculations

Uncertainty of the prediction (OECD principle 4 - Uncertainty of the prediction):
The prediction is based on 4 values, 4 of them (100%) equal to predicted value
Prediction confidence is measured by the p-value: 0.0123
Mechanistic interpretation:
The hypothesis for this prediction was built upon the identified mechanisms in the alerting profilers. In order to predict the potential for Ames mutagenicity of the query compound in S. thyphimurium TA 102 or E. Coli WP2 UvrA strains, the profiler alerts allowed for identification of similar substances which also share the same alerts within the general mechanistic "DNA binding by OECD" profiler, supported by corresponding alerts from the endpoint specific profiler "in vitro mutagenicity (Ames test) alerts by ISS", but the constraints on the source data restricted any identified compoudns to those only possessing data on these missing strains.
It is identified in the “Acylation >> Isocyanates and Isothiocyanates >> Isocyanates” alert in the general mechanistic profiler “DNA binding by OECD”, that the mechanism involves nucleophilic attack from biological nucleophiles.
Categorisation for the prediction was performed using the alerts discussed above first using the endpoint specific alert and then subcategorised with the general mechanistic alert also, this was followed by further subcategorization for chemical elements. This lead to a conclusion of negative for mutagenicity from the nearest neighbours (determined by log Kow) to the query structure which shared this categorisation. In each case, these isocyanates have results on either S. thyphimurium TA 102 or E. Coli WP2 UvrA.

Adequacy of the prediction:
Similar compounds sharing these alerts were identified, and using the nearest neighbours to the query compound, a prediction of negative for Ames mutagenicity was derived, using only data from S. thyphimurium TA 102 or E. Coli WP2 UvrA, lending weight to the already establish experimental result on the query compound in other strains.

Target profiles (OECD principle 5 - Chemical and biological mechanisms)
Profiles used for grouping/subcategorization Profile criteria:
- Acylation >> Isocyanates and Isothiocyanates >> Isocyanates (DNA binding by OECD) (primary grouping)
-- Target profiles: Acylation; Acylation >> Isocyanates and Isothiocyanates; Acylation >> Isocyanates and Isothiocyanates >> Isocyanates
- in vitro mutagenicity (Ames test) alerts by ISS (subcategorization)
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine, hydroxyl amine and its derived esters
- Organic functional groups (subcategorization)
-- Target profiles: Alkyl (hetero)arenes; Alkyl-, alkenyl- and alkynyl (hetero)arenes; Aryl; Isocyanate

Predefined Profile criteria
-OECD HPV Chemical Categories
-- Target profiles: Not categorized
-Substance type
-- Target profiles: Discrete chemical; Organic; Mono constituent (predefined)
-US-EPA New Chemical Categories
-- Target profiles: Not categorized

General Mechanistic
Profile criteria
-DNA binding by OECD
-- Target profiles: Acylation; Acylation >> Isocyanates and Isothiocyanates; Acylation >> Isocyanates and Isothiocyanates >> Isocyanates
-Protein binding by OASIS
-- Target profiles: Acylation; Acylation >> (Tio)carbamoylation of protein nucleophiles; Acylation >> (Tio)carbamoylation of protein nucleophiles >> Isothiocyanates, Isocyanates
Protein binding by OECD
-- Target profiles: Acylation; Acylation >> Isocyanates and Related Chemicals; Acylation >> Isocyanates and Related Chemicals >> Isocyanates
DNA binding by OASIS
-- Target profiles: No alert found

Endpoint Specific
- Aquatic toxicity classification by ECOSAR
-- Target profiles: Not Related to an Existing ECOSAR Class
- Protein binding alerts for Chromosomal aberration by OASIS
-- Target profiles: Acylation; Acylation >> (Thio)carbamoylation of protein nucleophiles; Acylation >> (Thio)carbamoylation of protein nucleophiles >> Isocyanates and Diisocyanates
-in vivo mutagenicity (Micronucleus) alerts by ISS
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine, hydroxyl amine and its derived esters
-DNA alerts for AMES, CA and MNT by OASIS
-- Target profiles: No alert found
-in vitro mutagenicity (Ames test) alerts by ISS
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine,hydroxyl amine and its derived esters

Empiric
Profile criteria
-Organic functional groups
-- Target profiles: Aryl; Isocyanate; Alkyl (hetero)arenes; Alkyl-, alkenyl- and alkynyl (hetero)arenes
-Organic functional groups (nested)
-- Target profiles: Isocyanate; Alkyl (hetero)arenes; Alkyl-, alkenyl- and alkynyl (hetero)arenes; Overlapped groups
-Organic functional groups (US EPA)
-- Target profiles: Isocyanate, aromatic attach [-N=C=O]; Miscellaneous sulfide (=S) or oxide (=O); Aliphatic Carbon [-CH3]; Aliphatic Carbon [-CH2-]; Aliphatic Carbon [CH]; Acetylenic Carbon [#C]; Aromatic Carbon [C]; Aliphatic Nitrogen, one aromatic attach [-N]
-Organic functional groups, Norbert Haider (checkmol)
-- Target profiles: CO2 derivative (general); Isocyanate; Aromatic compound
-Structure similarity
-- Target profiles: [90%,100%]
-Groups of elements
-- Target profiles: Non-Metals
-Lipinski Rule Oasis
-- Target profiles: Bioavailable
-Chemical elements
-- Target profiles: Group 14 - Carbon C; Group 15 - Nitrogen N; Group 16 - Oxygen O

log Kow (calculated): 3.14


Analogue(s) selection
(OECD principle 3 - Applicability domain)
Database(s) used:
- Bacterial mutagenicity ISSSTY
- Genotoxicity OASIS
- Toxicity Japan MHLW
Category boundaries (applicability domain):
- Active descriptor(s) range:
- log Kow: from 0.786 to 5.22 target chemical is in domain
- Response range: - E. Coli WP2 UvrA TA 102 W3102 E.coli WP2 PKM 101 E. Coli WP2 Uvr A WP2 Uvr A PKM 101 WP2 UvrA WP2 UvrA/pKM101: from Negative to Negative
Profilers:
- Acylation >> Isocyanates and Isothiocyanates >> Isocyanates (DNA binding by OECD) (primary grouping) target chemical is in domain
- in vitro mutagenicity (Ames test) alerts by ISS (subcategorization) target chemical is in domain
- Organic functional groups (subcategorization) target chemical is in domain
Additional data pruning: Cannot calculate X descriptor(s) 18 value(s) from 10 chemical(s)
Manually eliminated data points: none

Principles of method if other than guideline:

- Software tool(s) used including version: OECD QSAR Toolbox
- Model(s) used: v4.5
- Model description: see field 'Justification for non-standard information',
- Justification of QSAR prediction: see field 'Justification for type of information'

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

SMILES: Cc1cccc(c1)N=C=O

Species / strain / cell type:

S. typhimurium TA 102

Species / strain / cell type:

E. coli WP2

Key result

Species / strain:

other: E. coli WP2 and S. thyphimurium TA 102

Metabolic activation:

not applicable

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not applicable

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

not applicable

Remarks on result:

no mutagenic potential (based on QSAR/QSPR prediction)

Remarks:

Negative prediction for Ames mutagenicity via data exclusively from studies involving E.coli or TA 102 strains not covered by existing experimental Ames studies on the query compound.

Conclusions:

Ames mutagenicity via data exclusively from studies involving E.coli or TA 102 strains and inclusive of all required strains was predicted to be negative by the SAR.

Executive summary:

The E. coli WP2 and S. thyphimurium TA 102 strains have not been assessed for the query structure experimentally, however studies on other strains were shown to be negative.

In order to specifically assess activity towards E. coli and S. thyphimurium TA 102, it was determined that defining an endpoint in the OECD QSAR Toolbox with these strains specifically, cross referenced against any profiling alerts specific to Ames mutagenicity would allow for a robust prediction for the query structure.

 

The endpoint specific profiler "in vitro mutagenicity (Ames test) alerts by ISS" suggested Isocyanates undergo acylation directly without metabolic activation, involving reaction with DNA, where Isocyanates are shown to interact with cytosine, adenine and guanine. This was supported by the General mechanistic profiler “DNA binding by OECD”, which proposed that the mechanism involves nucleophilic attack from biological nucleophiles. This is of note as the mechanisms discussed around acylation via isocyanates would have been identified by the already studied strains i.e. the GC pair is present in these strains.

The prediction was performed using the alerts discussed above. This lead to a conclusion of negative for mutagenicity from the nearest neighbours (determined by log K_OW) to the query structure. In each case, these compounds have results on either S. thyphimurium TA 102 and/or E. coli WP2.

 

Based on the identified activities in profilers relevant to the Ames mutagenicity endpoint as discussed above and identifying those compounds which have available data in the missing strains, it is shown that the nearest neighbours to the test substance are all negative, therefore lending weight of evidence to the existing argument from the Ames study in the other strains that the test substance has not been tested on are also negative for Ames mutagenicity test.

Reference 2

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

(Q)SAR

Adequacy of study:

other information

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model, but not (completely) falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

1. SOFTWARE
OECD QSAR Toolbox

2. MODEL (incl. version number)
v4.5

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
Cc1cccc(c1)N=C=O

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
[Explain how the model fulfils the OECD principles for (Q)SAR model validation. Consider attaching the QMRF and/or QPRF or providing a link]
see TPRF information below

5. APPLICABILITY DOMAIN
[Explain how the substance falls within the applicability domain of the model]
see TPRF information below

6. ADEQUACY OF THE RESULT
see TPRF information below

Prediction summary
Predicted endpoint: Gene mutation; No effect specified; No species specified; No duration specified; No guideline specified
Predicted value: Negative
Unit/scale: Gene mutation I
Data gap filling method: Read-across analysis
Summary:
The Ames mutagenicity of m-tolylisocyanate was predicted for the strains S. thyphimurium TA 102 or E. Coli WP2 UvrA, which were not present in existing Ames studies on the query compound. The Toolbox endpoint was specifically defined as being Genetic Toxicity specific to the strains S. thyphimurium TA 102 or E. Coli WP2 UvrA, To derive the prediction, a combination of two alerts from one endpoint specific profiler was used to assess the identified aromatic amine mechanism related to the query compound.
Similar compounds sharing these alerts were identified, and using the nearest neighbours to the query compound, a prediction of negative for Ames mutagenicity was derived, using only data from S. thyphimurium TA 102 or E. Coli WP2 UvrA, lending weight to the already establish experimental result on the query compound in other strains.

Predicted value: Negative
Predicted endpoint (OECD Principle 1 - Defined endpoint): Human Health Hazards -> Genetic Toxicity ->Bacterial Reverse Mutation Assay (e.g. Ames Test) -> Gene mutation -> E. Coli WP2 UvrA TA 102 WP2 UvrA
Prediction plot: (see Figure 1 below)

Calculation approach (OECD principle 2 - Unambiguous algorithm):
takes the highest mode value from the 5 nearest neighbours
Active descriptor: log Kow (calculated) Data usage: Maximal value* *When multiple values are available for the same chemical, their maximal value is taken in prediction calculations

Uncertainty of the prediction (OECD principle 4 - Uncertainty of the prediction):
The prediction is based on 5 values, 5 of them (100%) equal to predicted value Prediction confidence is measured by the p-value: 0.00412
Mechanistic interpretation:
The hypothesis for this prediction was built upon the identified mechanisms in the alerting profilers. In order to predict the potential for Ames mutagenicity of the query compound in S. thyphimurium TA 102 or E. Coli WP2 UvrA strains, the profiler alerts allowed for identification of similar substances which also share the aromatic amines alert from the endpoint specific profiler " in vitro mutagenicity (Ames test) alerts by ISS " profiler, but the constraints on the source data restricted any identified compounds to those only possessing data on these missing strains. The identified mechanisms for the action of aromatic amines is discussed in both the isocyanate alert and the Primary Aromatic amine alert. With the former suggesting that aromatic isocyanates specifically can undergo hydrolysis to carbamic acid and subsequent decarboxylation to the corresponding aromatic amine. The specific alert for aromatic amines further discusses the metabolic pathway for these compounds to elicit covalent bonding to DNA. The compounds are proposed to undergo cytochrome P450 mediated N-oxidation to N-hydroxyarylamines, which are then further transformed by enzymatic esterification to highly reactive nitrenium ion species which can bind covalently to biological molecules to form aminoaryl derivatives. Using the alerts discussed above, the prediction was able to derive a conclusion of negative from the nearest neighbours to the test substance structure. In each case, these aromatic amines have results on either S. thyphimurium TA 102 or E. Coli WP2 UvrA. From the below it can be determined that for the specific strains the nearest neighbours containing tertiary amines, which are analogous with the specific moiety of the test substance, all results in these strains are negative, which is in concordance with the existing study results available for the target substance on other strains.
Adequacy of the prediction:
All assessment criteria were within domain. Similar compounds sharing these alerts were identified, and using the nearest neighbours to the query compound, a prediction of negative for Ames mutagenicity was derived, using only data from S. thyphimurium TA 102 or E. Coli WP2 UvrA, lending weight to the already establish experimental result on the query compound in other strains.

Target profiles (OECD principle 5 - Chemical and biological mechanisms)
Profiles used for grouping/subcategorization Profile criteria:
-Primary aromatic amine,hydroxyl amine and its derived esters (in vitro mutagenicity (Ames test) alerts by ISS) (primary grouping)
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine,hydroxyl amine and its derived esters
-in vitro mutagenicity (Ames test) alerts by ISS (subcategorization)
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine,hydroxyl amine and its derived esters
-Chemical elements (subcategorization)
-- Target profiles: Group 14 - Carbon C; Group 15 - Nitrogen N; Group 16 - Oxygen O
-Structure similarity (subcategorization)
-- Target profiles: [90%,100%]

Predefined Profile criteria
-OECD HPV Chemical Categories
-- Target profiles: Not categorized
-Substance type
-- Target profiles: Discrete chemical; Organic; Mono constituent (predefined)
-US-EPA New Chemical Categories
-- Target profiles: Not categorized

General Mechanistic
Profile criteria
-DNA binding by OECD
-- Target profiles: Acylation; Acylation >> Isocyanates and Isothiocyanates; Acylation >> Isocyanates and Isothiocyanates >> Isocyanates
-Protein binding by OASIS
-- Target profiles: Acylation; Acylation >> (Tio)carbamoylation of protein nucleophiles; Acylation >> (Tio)carbamoylation of protein nucleophiles >> Isothiocyanates, Isocyanates
Protein binding by OECD
-- Target profiles: Acylation; Acylation >> Isocyanates and Related Chemicals; Acylation >> Isocyanates and Related Chemicals >> Isocyanates
DNA binding by OASIS
-- Target profiles: No alert found

Endpoint Specific
- Aquatic toxicity classification by ECOSAR
-- Target profiles: Not Related to an Existing ECOSAR Class
- Protein binding alerts for Chromosomal aberration by OASIS
-- Target profiles: Acylation; Acylation >> (Thio)carbamoylation of protein nucleophiles; Acylation >> (Thio)carbamoylation of protein nucleophiles >> Isocyanates and Diisocyanates
-in vivo mutagenicity (Micronucleus) alerts by ISS
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine, hydroxyl amine and its derived esters
-DNA alerts for AMES, CA and MNT by OASIS
-- Target profiles: No alert found
-in vitro mutagenicity (Ames test) alerts by ISS
-- Target profiles: Isocyanate and isothiocyanate groups; Primary aromatic amine,hydroxyl amine and its derived esters

Empiric
Profile criteria
-Organic functional groups
-- Target profiles: Aryl; Isocyanate; Alkyl (hetero)arenes; Alkyl-, alkenyl- and alkynyl (hetero)arenes
-Organic functional groups (nested)
-- Target profiles: Isocyanate; Alkyl (hetero)arenes; Alkyl-, alkenyl- and alkynyl (hetero)arenes; Overlapped groups
-Organic functional groups (US EPA)
-- Target profiles: Isocyanate, aromatic attach [-N=C=O]; Miscellaneous sulfide (=S) or oxide (=O); Aliphatic Carbon [-CH3]; Aliphatic Carbon [-CH2-]; Aliphatic Carbon [CH]; Acetylenic Carbon [#C]; Aromatic Carbon [C]; Aliphatic Nitrogen, one aromatic attach [-N]
-Organic functional groups, Norbert Haider (checkmol)
-- Target profiles: CO2 derivative (general); Isocyanate; Aromatic compound
-Structure similarity
-- Target profiles: [90%,100%]
-Groups of elements
-- Target profiles: Non-Metals
-Lipinski Rule Oasis
-- Target profiles: Bioavailable
-Chemical elements
-- Target profiles: Group 14 - Carbon C; Group 15 - Nitrogen N; Group 16 - Oxygen O

log Kow (calculated): 3.14


Analogue(s) selection
(OECD principle 3 - Applicability domain)
Database(s) used:
- Bacterial mutagenicity ISSSTY
- Genotoxicity OASIS
- Toxicity Japan MHLW
Category boundaries (applicability domain):
- Active descriptor(s) range:
- log Kow: from 1.62 to 3.74 target chemical is in domain
- Response range: - E. Coli WP2 UvrA TA 102 W3102 E.coli WP2 PKM 101 E. Coli WP2 Uvr A WP2 Uvr A PKM 101 WP2 UvrA WP2 UvrA/pKM101: from Negative to Negative
Profilers:
- Primary aromatic amine,hydroxyl amine and its derived esters (in vitro mutagenicity (Ames test) alerts by ISS) (primary grouping) target chemical is in domain
- in vitro mutagenicity (Ames test) alerts by ISS (subcategorization) target chemical is in domain - Chemical elements (subcategorization) target chemical is in domain
- Structure similarity (subcategorization) target chemical is in domain
· Mode: Hologram, CombineAllFeatures
· Measure: Dice
· Molecular features: AtomCenteredFragments
· Atom characteristics: AtomType,
CountHAttached, Hybridization
· Differ from target by: At least one category
· Removed categories: [0%,10%); [10%,20%); [20%,30%); [30%,40%)
Additional data pruning: Cannot calculate X descriptor(s) 18 value(s) from 10 chemical(s)
Manually eliminated data points: none

Principles of method if other than guideline:

- Software tool(s) used including version: OECD QSAR Toolbox
- Model(s) used: v4.5
- Model description: see field 'Justification for non-standard information',
- Justification of QSAR prediction: see field 'Justification for type of information'

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

SMILES: Cc1cccc(c1)N=C=O

Species / strain / cell type:

S. typhimurium TA 102

Species / strain / cell type:

E. coli WP2

Key result

Species / strain:

other: E. coli WP2 and S. thyphimurium TA 102

Metabolic activation:

not applicable

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not applicable

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

not applicable

Remarks on result:

no mutagenic potential (based on QSAR/QSPR prediction)

Remarks:

Negative prediction for Ames mutagenicity via data exclusively from studies involving E.coli or TA 102 strains not covered by existing experimental Ames studies on the query compound.

Conclusions:

Ames mutagenicity via data exclusively from studies involving E.coli or TA 102 strains and inclusive of all required strains was predicted to be negative by the SAR.

Executive summary:

The E. coli WP2 and S. thyphimurium TA 102 strains have not been assessed for the query structure experimentally, however studies on other strains were shown to be negative.

In order to specifically assess activity towards E. coli and S. thyphimurium TA 102, it was determined that defining an endpoint in the OECD QSAR Toolbox with these strains specifically, cross referenced against any profiling alerts specific to Ames mutagenicity would allow for a robust prediction for the query structure.

 

The endpoint specific profiler "in vitro mutagenicity (Ames test) alerts by ISS" suggested the query structure may elicit covalent bonding to DNA, by first undergoing hydrolysis to carbamic acid and subsequent decarboxylation to the corresponding aromatic amine, after which aromatic amines are proposed to undergo cytochrome P450 mediated N-oxidation to N-hydroxyarylamines, which are then further transformed by enzymatic esterification to highly rection nitrenium ion species which can bind covalently to biological molecules to form aminoaryl derivatives. 

The prediction was performed using the alerts discussed above. This lead to a conclusion of negative for mutagenicity from the nearest neighbours (determined by log K_OW) to the query structure. In each case, these compounds have results on either S. thyphimurium TA 102 and/or E. coli WP2.

 

Based on the identified activities in profilers relevant to the Ames mutagenicity endpoint as discussed above and identifying those compounds which have available data in the missing strains, it is shown that the nearest neighbours to the test substance are all negative, therefore lending weight of evidence to the existing argument from the Ames study in the other strains that the test substance has not been tested on are also negative for Ames mutagenicity test.

Reference 3

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

other information

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: guideline study - only 4 strains tested

Justification for type of information:

Added QSAR in silico prediction for E. coli WP2 and S. thyphimurium TA 102 strains. (WoE,BFS, 2022, AMES QSAR, Aromatic Amines and WoE,BFS, 2022, AMES QSAR, Isocyanates)

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

The mutagenic potential of m-tolylisocyanate was examined in the Salmonella/microsome test. Bacteria of four histidineauxotrophic Salmonella typhimurium LT2 mutant strains (TA 98, TA 100, TA 1535, and TA 1537) were exposed to doses up to 5000 pg per plate. Both the plate incorporation method and the preincubation method was used.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

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

Additional strain / cell type characteristics:

other: Firstly, they are deep rough since certain lipopolysaccharide side chains are missing in the bacterial cell wall. Secondly, their reduced ability to repair damage from UV light (e.g. thymidine dimers) allows the phenotypic detection of mutation events

Metabolic activation:

with and without

Metabolic activation system:

S-9 mix

Test concentrations with justification for top dose:

plate incorporation assay: 0, 8, 40, 200, 1000, 5000 µg/plate
preincubation assay: 0, 62.5, 125, 250, 500, 1000, 2000 µg/tube

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

other: sodium azide, nitrofurantoin, 4-nitro- 1,2-phenylene diamine, 2-aminoanthracene

Details on test system and experimental conditions:

IUCLID4 Type: Ames test

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

other: Doses up to and including 200 pg per plate did not cause any bacteriotoxic effects

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

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

Remarks:

Migrated from field 'Test system'.

Doses up to and including 200 µg per plate did not cause any bacteriotoxic effects: Total bacteria counts remained unchanged and no growth inhibition was observed. The substance revealed weak, strain-specific bacteriotoxic effects at higher doses yet doses up to 1000 µg per plate could still be used for assessment purposes. Substance precipitation occurred at the dose of 500 µg per plate and above. Plates containing 2000 or 5000 µg of the test substance could not be used for assessment purposes.

Conclusions:

Interpretation of results (migrated information):
negative

Executive summary:

The mutagenic potential of m-tolylisocyanate was examined in the Salmonella/microsome test. Bacteria of four histidineauxotrophic Salmonella typhimurium LT2 mutant strains (TA 98, TA 100, TA 1535, and TA 1537) were exposed to doses up to 5000 pg per plate. Both the plate incorporation method and the preincubation method was used.

Doses up to and including 200 µg per plate did not cause any bacteriotoxic effects: Total bacteria counts remained unchanged and no growth inhibition was observed. The substance revealed weak, strain-specific bacteriotoxic effects at higher doses yet doses up to 1000 µg per plate could still be used for assessment purposes. Substance precipitation occurred at the dose of 500 µg per plate and above. Plates containing 2000 or 5000 µg of the test substance could not be used for assessment purposes.

There was no evidence for mutagenic effects of m-tolylisocyanate with and without S9 mix. A biologically relevant increase of revertant colony numbers over control levels was not observed. Therefore, m-tolylisocyanate was considered to be non-mutagenic in the Salmonella/microsome test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

In a valid Ames test m-tolyl isocyanante was negative

Further QSAR assessment was conducted to consider E. coli WP2 and S. thyphimurium TA 102 strains, which also concluded negative
Short description of key information:

in vitro:
Ames test: negative
Cytogenetic assay: no data
mammalian cell gene mutation test: no data

The mutagenic potential of m-tolylisocyanate was examined in the Salmonella/microsome test. Bacteria of four histidineauxotrophic Salmonella typhimurium LT2 mutant strains (TA 98, TA 100, TA 1535, and TA 1537) were exposed to doses up to 5000 pg per plate. Both the plate incorporation method and the preincubation method was used.

Doses up to and including 200 µg per plate did not cause any bacteriotoxic effects: Total bacteria counts remained unchanged and no growth inhibition was observed. The substance revealed weak, strain-specific bacteriotoxic effects at higher doses yet doses up to 1000 µg per plate could still be used for assessment purposes. Substance precipitation occurred at the dose of 500 µg per plate and above. Plates containing 2000 or 5000 µg of the test substance could not be used for assessment purposes.

There was no evidence for mutagenic effects of m-tolylisocyanate with and without S9 mix. A biologically relevant increase of revertant colony numbers over control levels was not observed. Therefore, m-tolylisocyanate was considered to be non-mutagenic in the Salmonella/microsome test.

in silico:

Information valid as supporting data.

Ames mutagenicity: negative

Aromatic Amines:

The E. coli WP2 and S. thyphimurium TA 102 strains have not been assessed for the query structure experimentally, however studies on other strains were shown to be negative.

In order to specifically assess activity towards E. coli and S. thyphimurium TA 102, it was determined that defining an endpoint in the OECD QSAR Toolbox with these strains specifically, cross referenced against any profiling alerts specific to Ames mutagenicity would allow for a robust prediction for the query structure.

 

The endpoint specific profiler "in vitro mutagenicity (Ames test) alerts by ISS" suggested the query structure may elicit covalent bonding to DNA, by first undergoing hydrolysis to carbamic acid and subsequent decarboxylation to the corresponding aromatic amine, after which aromatic amines are proposed to undergo cytochrome P450 mediated N-oxidation to N-hydroxyarylamines, which are then further transformed by enzymatic esterification to highly rection nitrenium ion species which can bind covalently to biological molecules to form aminoaryl derivatives. 

The prediction was performed using the alerts discussed above. This lead to a conclusion of negative for mutagenicity from the nearest neighbours (determined by log K_OW) to the query structure. In each case, these compounds have results on either S. thyphimurium TA 102 and/or E. coli WP2.

 

Based on the identified activities in profilers relevant to the Ames mutagenicity endpoint as discussed above and identifying those compounds which have available data in the missing strains, it is shown that the nearest neighbours to the test substance are all negative, therefore lending weight of evidence to the existing argument from the Ames study in the other strains that the test substance has not been tested on are also negative for Ames mutagenicity test.

 

Isocyanates:

The E. coli WP2 and S. thyphimurium TA 102 strains have not been assessed for the query structure experimentally, however studies on other strains were shown to be negative.

In order to specifically assess activity towards E. coli and S. thyphimurium TA 102, it was determined that defining an endpoint in the OECD QSAR Toolbox with these strains specifically, cross referenced against any profiling alerts specific to Ames mutagenicity would allow for a robust prediction for the query structure.

 

The endpoint specific profiler "in vitro mutagenicity (Ames test) alerts by ISS" suggested Isocyanates undergo acylation directly without metabolic activation, involving reaction with DNA, where Isocyanates are shown to interact with cytosine, adenine and guanine. This was supported by the General mechanistic profiler “DNA binding by OECD”, which proposed that the mechanism involves nucleophilic attack from biological nucleophiles. This is of note as the mechanisms discussed around acylation via isocyanates would have been identified by the already studied strains i.e. the GC pair is present in these strains.

The prediction was performed using the alerts discussed above. This lead to a conclusion of negative for mutagenicity from the nearest neighbours (determined by log K_OW) to the query structure. In each case, these compounds have results on either S. thyphimurium TA 102 and/or E. coli WP2.

 

Based on the identified activities in profilers relevant to the Ames mutagenicity endpoint as discussed above and identifying those compounds which have available data in the missing strains, it is shown that the nearest neighbours to the test substance are all negative, therefore lending weight of evidence to the existing argument from the Ames study in the other strains that the test substance has not been tested on are also negative for Ames mutagenicity test.

 

in vivo:
no data

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Harmonised classification:

The substance does not have a harmonised classification according to the Regulation (EC) No. 1272/2008 (CLP). 

 

Self-classification:

Based on the available information on the registered substance, self-classification is proposed according to GHS and CLP:

GHS not classified.