Registration Dossier

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The results of the Bacterial Reverse Mutation Assay indicate that, under the conditions of this study, OFPMA did not cause a positive mutagenic response with any of the tester strains in either the presence or absence of Aroclor-induced rat liver S9.

 

The test article, OFPMA, was considered negative for inducing micronuclei in binucleated cells in cultured human lymphocytes without and with metabolic activation.

 

The in vitro gene mutation study in mammalian cells has been waived based on consideration of the results of the available studies, the lack of structural alters for genotoxicity or for carcinogenicity in the substance and its expected metabolites, and according to EFSA guidance, should a compound be found to be without activity in the in vitro gene mutation study in bacteria and in the in vitro micronucleus assay, no further in vitro or in vivo testing is considered necessary. Therefore, the in vitro mammalian cell test will not provide any further useful information about the potential in vivo mutagenicity of OFPMA.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05 February 2016 to 22 February 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1997
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
Purity: 99.7%
Target gene:
selected loci of several strains of Salmonella typhimurium and at the tryptophan locus of Escherichia coli strain WP2 uvrA
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Details on mammalian cell type (if applicable):
not applicable
Additional strain / cell type characteristics:
not applicable
Cytokinesis block (if used):
not applicable
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254-induced rat liver S9 was used as the metabolic activation system.
Test concentrations with justification for top dose:
Concentration levels of 1.50, 5.00, 15.0, 50.0, 150, 500, 1500 and 5000 μg per plate in DMSO (highest exposure concentration per OECD test guideline 471).

The initial toxicity-mutation assay was used to establish the dose-range for the confirmatory mutagenicity assay and to provide a preliminary mutagenicity evaluation. TA98, TA100, TA1535, TA1537 and WP2 uvrA were exposed to the vehicle alone, positive controls and eight dose levels of the test article, in duplicate, in the presence and absence of S9-mix. Dose levels for the confirmatory mutagenicity assay were based upon absence of post-treatment toxicity.

The confirmatory mutagenicity assay was used to evaluate and confirm the mutagenic potential of the test article. TA98, TA100, TA1535, TA1537 and WP2 uvrA were exposed to the vehicle alone, positive controls and six dose levels of the test article, in triplicate, in the presence and absence of S9-mix.
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: based on the information provided by the Sponsor and the solubility of the test article and compatibility with the target cells.
Untreated negative controls:
yes
Negative solvent / vehicle controls:
no
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: 2-aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation) for Experiment 1; preincubation for Experiment 2

NUMBER OF REPLICATIONS: duplicates for Experiment 1; triplicates for Experiment 2

Criteria for a Valid Test
The following criteria must be met for each assay to be considered valid:
All Salmonella tester strain cultures must demonstrate the presence of the deep rough mutation (rfa) and the deletion in the uvrB gene. Cultures of tester strains TA98 and TA100 must demonstrate the presence of the pKM101 plasmid R-factor. All WP2 uvrA cultures must demonstrate the deletion in the uvrA gene.
All cultures must demonstrate the characteristic mean number of spontaneous revertants in the vehicle controls as follows (inclusive): TA98, 10 - 50; TA100, 80 - 240; TA1535, 5 - 45; TA1537, 3 - 21; WP2 uvrA, 10 - 60.

To ensure that appropriate numbers of bacteria are plated, tester strain culture titers must be greater than or equal to 0.3x109 cells/mL.

The mean of each positive control must exhibit at least a 3.0-fold increase in the number of revertants over the mean value of the respective vehicle control.

A minimum of three non-toxic dose levels is required to evaluate assay data. A dose level is considered toxic if one or both of the following criteria are met:
(1) A >50 % reduction in the mean number of revertants per plate as compared to the mean vehicle control value. This reduction must be accompanied by an abrupt dose-dependent drop in the revertant count.
(2) At least a moderate reduction in the background lawn (background code 3, 4 or 5).
Rationale for test conditions:
Results from the plate incorporation method were clarified by further testing using the preincubation method.
Evaluation criteria:
The condition of the bacterial background lawn was evaluated for evidence of test article toxicity by using a dissecting microscope. Precipitate was evaluated after the incubation period by visual examination without magnification. Toxicity and degree of precipitation were scored relative to the vehicle control plate. As appropriate, colonies were enumerated either by hand or by machine.

For each replicate plating, the mean and standard deviation of the number of revertants per plate were calculated and are reported.

For the test article to be evaluated positive, it must cause a dose-related increase in the mean revertants per plate of at least one tester strain over a minimum of two increasing concentrations of test article as specified below:

Strains TA1535 and TA1537
Data sets were judged positive if the increase in mean revertants at the peak of the dose response was equal to or greater than 3.0-times the mean vehicle control value.

Strains TA98, TA100 and WP2 uvrA
Data sets were judged positive if the increase in mean revertants at the peak of the dose response was equal to or greater than 2.0-times the mean vehicle control value.

An equivocal response is a biologically relevant increase in a revertant count that partially meets the criteria for evaluation as positive. This could be a dose-responsive increase that does not achieve the respective threshold cited above or a non-dose responsive increase that is equal to or greater than the respective threshold cited. A response was evaluated as negative if it was neither positive nor equivocal.
Statistics:
The mean values and standard deviation were calculated for the number of revertant colony count.
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
toxicity was observed beginning at 500 μg per plate without metabolic activation
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
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
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:
cytotoxicity
Remarks:
toxicity was observed beginning at 500 μg per plate without metabolic activation
Untreated negative controls validity:
valid
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
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Precipitation was not observed in either test 1 or 2. Cytotoxicity was not observed in the absence or presence of metabolic activation in Experiments 1 and 2.

No positive mutagenic responses were observed in the presence or absence of metabolic activation in any of the tested strains in either Experiment 1 or 2.
Conclusions:
All criteria for a valid study were met as described in the protocol. The results of the Bacterial Reverse Mutation Assay indicate that, under the conditions of this study, the test material did not cause a positive mutagenic response with any of the tester strains in either the presence or absence of Aroclor-induced rat liver S9.
Executive summary:

The test article was tested to evaluate its mutagenic potential by measuring its ability to induce reverse mutations at selected loci of several strains of Salmonella typhimurium and at the tryptophan locus of Escherichia coli strain WP2 uvrA in the presence and absence of an exogenous metabolic activation system. Dimethyl sulfoxide (DMSO) was used as the vehicle.

In the initial toxicity-mutation assay via plate incorporation method, the dose levels tested were 1.50, 5.00, 15.0, 50.0, 150, 500, 1500 and 5000 μg per plate. Neither precipitate nor toxicity was observed. No positive mutagenic responses were observed with any of the tester strains in either the presence or absence of S9 activation. Based upon these results, the maximum dose tested in the confirmatory mutagenicity assay was 5000 μg per plate.

In the confirmatory mutagenicity assay via pre-incubation method, the dose levels tested were 15.0, 50.0, 150, 500, 1500 and 5000 μg per plate. No precipitate was observed. Toxicity was observed beginning at 500 or at 5000 μg per plate with tester strains TA100 and TA1535 in the absence of S9 activation. No positive mutagenic responses were observed with any of the tester strains in either the presence or absence of S9 activation.

These results indicate OFPMA was negative for the ability to induce reverse mutations at selected loci of several strains of Salmonella typhimurium and at the tryptophan locus of Escherichia coli strain WP2 uvrA in the presence and absence of an exogenous metabolic activation system.

Endpoint:
in vitro cytogenicity / micronucleus study
Type of information:
experimental study
Adequacy of study:
key study
Study period:
14 July 2008 to 13 October 2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 487 (In vitro Mammalian Cell Micronucleus Test)
Version / remarks:
2008
Deviations:
no
GLP compliance:
yes
Type of assay:
in vitro mammalian cell micronucleus test
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Lot No. Q151-170
- purity: 99%
Species / strain / cell type:
lymphocytes:
Details on mammalian cell type (if applicable):
CELLS USED
- Source of cells: human peripheral blood lymphocytes
Metabolic activation:
with and without
Metabolic activation system:
Aroclor™ 1254 induced rat liver post-mitochondrial fraction (S9) coupled with NADP and isocitric acid
Test concentrations with justification for top dose:
Without metabolic activation:
Test 1 (exposure - 3 h exposure; harvest time - 24 h): 0, 250, 500, 1000 μg/mL
Test 2 (exposure - 24 h exposure; harvest time - 24 h): 0, 125, 250, 500, 1000 μg/mL
Test 3 (exposure - 3 h exposure; harvest time - 24 h): 0, 250, 400, 500, 750, 1000 μg/mL
Test 4 (exposure - 24 h exposure; harvest time - 24 h): 0, 62.5, 125, 175, 250, 300, 400, 500, 750, 1000 μg/mL
Test 5 (exposure - 3 h exposure; harvest time - 24 h): 0*, 300, 400, 500, 525, 550, 575, 600, 625, 650*, 700*, 750, 800* μg/mL
Test 6 (exposure - 24 h exposure; harvest time - 24 h): 0*, 150, 175, 200, 213*, 225*, 250*, 275, 300, 400 μg/mL
With metabolic activation:
Test 1 (exposure - 3 h exposure; harvest time - 24 h): 0, 250, 500, 1000 μg/mL
Test 2 (exposure - 3 h exposure; harvest time - 24 h): 0*, 400, 500*, 750*, 1000* μg/mL
*Cultures selected for metaphase analysis
Top test concentrations were selected based on cytotoxicity.
Vehicle / solvent:
Dimethylsulfoxide (DMSO)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
mitomycin C
Details on test system and experimental conditions:
Without metabolic activation:
Test 1: exposure - 3 h exposure; harvest time - 24 h
Test 2: exposure - 24 h exposure; harvest time - 24 h
Test 3: exposure - 3 h exposure; harvest time - 24 h
Test 4: exposure - 24 h exposure; harvest time - 24 h
Test 5: exposure - 3 h exposure; harvest time - 24 h
Test 6: exposure - 24 h exposure; harvest time - 24 h
With metabolic activation:
Test 1: exposure - 3 h exposure; harvest time - 24 h
Test 2: exposure - 3 h exposure; harvest time - 24 h

Cytochalasin B was used to block the cytoplasmic cell division after the treatment in order to observe the nuclear status of the test cells.
Rationale for test conditions:
The assay is designed to establish whether the test article(s) or metabolites can interact with cells to inhibit the cell cycle and/or induce micronuclei.
Evaluation criteria:
The frequency BN-MN for the vehicle control cultures must be within historical ranges. The positive control articles must induce a statistically significant increase (p < 0.05) in the frequency of MN-BN, as compared to the concurrent vehicle controls.
Statistics:
Statistical analysis employed a Fisher-Irwin exact test for pair wise comparisons between each treated and vehicle control cultures. One-sided tail probabilities will be used to evaluate statistical significance (p < 0.05). A chemical was concluded to be positive if a statistically significant increase in the frequency of micronucleated cells was obtained from at least one test article concentration compared to the concurrent vehicle control and the linear trend test confirmed a positive dose-response (alpha level = 0.05).
Key result
Species / strain:
lymphocytes: human peripheral blood lymphocytes
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
Due to a lack of appropriate toxicity, the assays without metabolic activation were repeated.
Vehicle controls validity:
not applicable
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
No toxicologically significant increases in the number of cells with aberrations were noted, with or without metabolic activation. The positive and vehicle controls gave satisfactory responses confirming the validity of the test system.
Conclusions:
The notified chemical was not clastogenic to human peripheral blood lymphocytes treated in vitro under the conditions of the test.
Executive summary:

The objective of this in vitro assay was to evaluate the ability of OFPMA to cause micronuclei in cultured human lymphocytes using cytochalasin B-block methodology with and without an exogenous metabolic activation system.

 

Dimethylsulfoxide (DMSO) was the vehicle for the study. The highest concentration tested in the assay was 1000 μg/mL, which was above the solubility limit of the formulated OFPMA after dosing into culture medium. The stock formulations and their dilutions were dosed using a dosing volume of 1% (10.0 μL/mL) and the vehicle control cultures were treated with 10.0 μL/mL of DMSO.

 

In the dose range-finding study, the treatment periods were for 3 and ~24 hours without metabolic activation and 3 hours with metabolic activation. Cultures were harvested24 hours from the initiation of treatment. Concentrations of OFPMA ranging from 7.81 to 1000μg/mL were tested without and with metabolic activation. Based on observed cytotoxicity, doses were selected for the micronucleus assay.

 

In the micronucleus assay, the treatment period was for 3 and ~24 hours without metabolic activation and 3 hours with metabolic activation. Cultures were harvested ~24 hours from the initiation of treatment. The micronucleus assay was conducted testing concentrations of 125, 250, 400, 500, 750, and 1000 μg/mL without metabolic activation with a 3-hour treatment, at 62.5, 125, 175, 250, 300, 350, 400, 500, 750, and 1000 μg/mL without metabolic activation with a ~24-hour treatment, and at 125, 250, 400, 500, 750, and 1000 μg/mL with metabolic activation. Cultures treated with concentrations of 500, 750, and 1000 μg/mL with metabolic activation were analyzed for micronucleated binulceated cells. No significant increase in micronucleated binulceated cells was observed in the cultures analyzed. Due to a lack of appropriate toxicity, the assays without metabolic activation were repeated.

 

The repeat of the micronucleus assay was conducted testing concentrations of 300, 400, 500, 525, 550, 575, 600, 625, 650, 700, 750, and 800 μg/mL without metabolic activation with a 3-hour treatment, and at 50.0, 100, 150, 175, 200, 213, 225, 250, 275, 300, and 400 μg/mL without metabolic activation with a ~24-hour treatment. Cultures treated with concentrations of 650, 700, and 800 μg/mL without metabolic activation (3-hour treatment), and 213, 225, and 250 μg/mL without metabolic activation (~24-hour treatment) were analyzed for micronucleated binulceated cells. No significant increase in micronucleated binulceated cells was observed in the cultures analyzed.

 

The positive control cultures had significant increase in micronucleated binulceated cells as compared with the vehicle control cultures. The high doses selected for analysis in the assay had a precipitate at the end of the treatment period or ~50% reduction in CBPI as recommended for this assay by the OECD Testing Guidelines.

 

The test article was considered negative for inducing micronuclei in binulceated cells in cultured human lymphocytes without and with metabolic activation.

Endpoint:
in vitro gene mutation study in mammalian cells
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Justification for type of information:
An in vitro gene mutation study in mammalian cells is the second part of the standard information set required for registration at the Annex VIII tonnage level. ECHA’s Guidance on Information Requirements and Chemical Safety Assessment (ECHA, 2017) states it is possible to omit the second in vitro study in mammalian cells. If it can be demonstrated that this mammalian cell test will not provide any further useful information about the potential in vivo mutagenicity of a substance, then it does not need to be conducted.

Experimental data with OFPMA involving two in vitro gene mutation studies in bacteria (BioReliance Corporation, 2016; IIT Research Institute, 2008) and one in vitro micronucleus study (Covance Laboratories, 2009) are all negative; details are provided elsewhere within this dossier. This demonstrates that OFPMA has no genotoxixc potential and no further testing should be necessary. This is consistent with the fact that the in vitro gene mutation study in bacteria (Ames assay) (OECD TG 471) and the in vitro micronucleus study (OECD TG 487) are the 2 in vitro genotoxicity tests recommended by the European Food Safety Authority for initial assessment of genotoxic potential of food additives or ingredients. As stated in their guidance document (EFSA, 2012), should a compound be found to be without activity in the in vitro gene mutation study in bacteria and in the in vitro micronucleus assay, no further in vitro or in vivo testing is considered necessary. The combination of the Ames and in vitro micronucleus tests has been demonstrated to be sufficient to detect almost all in vivo genotoxic and/or carcinogenic compounds and that addition of a gene mutation assay in mammalian cells (i.e., mouse lymphoma assay) adds little value in this regard (Kirkland et al., 2011).
An examination of the structure of the notified substance clearly shows that the substance comprises a short-chained polyfluoroalkyl methacrylate. Assessment of the chemical structure of OFPMA has determined that the substance does not contain any structural alerts that are indicative of mutagenic, genotoxic or carcinogenic potential (Ashby and Tennant, 1991; Tennant and Ashby, 1991; Ashby, 1996; Benigni and Bossa, 2008).

The substance is anticipated to be metabolized to octafluoropentanol and methacrylic acid. The methacrylic acid is subsequently converted to carbon dioxide via the tricarboxylic acid cycle in both experimental animals and humans (WHO, 1998). The octafluoropentanol is expected to be converted to octafluoropentanal via alcohol dehydrogenase and octafluoropentanoic acid via aldehyde dehydrogenase. Assessment of the chemical structures of these metabolites likewise determined that the substances do not contain any structural alerts that are indicative of mutagenic, genotoxic or carcinogenic potential.

None of the chemical groups/moieties in the substance and its metabolites have been indicated as a structural alert for genotoxicity or for carcinogenicity. Therefore, the in vitro mammalian cell test will not provide any further useful information about the potential in vivo mutagenicity of OFPMA.

REFERENCES
Ashby J (1996). Alternatives to the 2-species bioassay for the identification of potential human carcinogens. Human Exp Toxicol 15:183-202.

Ashby J, Tennant RW (1991). Definitive relationships among chemical structure, carcinogenicity and mutagenicity for 301 chemicals tested by the U.S. NTP (National Toxicology Program). Mutat Res 257(3):229-306.

Benigni R, Bossa C (2008). Structure alerts for carcinogenicity, and the Salmonella assay system: a novel insight through the chemical relational databases technology. Mutat Res 659(3):248-261

BioReliance Corporation (2016). Bacterial Reverse Mutation Assay. BioReliance Corporation, 9630 Medical Center Drive, Rockville, MD 20850. Unpublished report number AE46LA.503005.BTL (provided elsewhere within this dossier)

Covance Laboratories (2009). In Vitro Human Peripheral Blood Lymphocyte Micronucleus Assay. Covance Laboratories Inc., 9200 Leesburg Pike, Vienna, Virginia 22182-1699. Unpublished report number 8097-100. (provided elsewhere within this dossier)

ECHA (2017). Guidance on information requirements and chemical safety assessment. Chapter R.7a: Endpoint specific guidance. Volume 3.0, July 2017. Available at: https://echa.europa.eu/documents/10162/13632/information_requirements_r7a_en.pdf/e4a2a18f-a2bd-4a04-ac6d-0ea425b2567f

EFSA (2012). Scientific Opinion. Scientific opinion on genotoxicity testing strategies applicable to food and feed safety assessment. EFSA Scientific Committee, European Food Safety Authority (EFSA), Parma, Italy. EFSA Journal 9(9):2379. Available at: http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2011.2379/epdf

IIT Research Institute (2008). Evaluation of the Potential Mutagenic Activity of OFPMA in the Bacterial Reverse Mutation Assay. IIT Research Institute, Life Sciences Group, 10 West 35th Street, Chicago, IL 60616-3799. Unpublished report number 2171-002. (provided elsewhere within this dossier)

Kirkland D, Reeve L, Gatehouse D, Vanparysd P (2011). A core in vitro genotoxicity battery comprising the Ames test plus the in vitro micronucleus test is sufficient to detect rodent carcinogens and in vivo genotoxins. Mutat Res 721(1):27-73

Tennant RW, Ashby J (1991). Classification according to chemical structure, mutagenicity to Salmonella and level of carcinogenicity of a further 39 chemicals tested for carcinogenicity by the U.S. National Toxicology Program. Mutat Res 257(3):209-227

WHO (1998). Concise International chemical assessment document 4. Methyl methacrylate. Geneva, Switzerland: World Health Organization (WHO). Available at: http://www.who.int/ipcs/publications/cicad/en/cicad04.pdf
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

AMES

The test article was tested to evaluate its mutagenic potential by measuring its ability to induce reverse mutations at selected loci of several strains of Salmonella typhimurium and at the tryptophan locus of Escherichia coli strain WP2 uvrA in the presence and absence of an exogenous metabolic activation system with an independent repeat study. Dimethyl sulfoxide (DMSO) was used as the vehicle.

 

These results indicate OFPMA was negative for the ability to induce reverse mutations at selected loci of several strains of Salmonella typhimurium and at the tryptophan locus of Escherichia coli strain WP2 uvrA in the presence and absence of an exogenous metabolic activation system.

 

In Vitro Micronucleus

The objective of this in vitro assay was to evaluate the ability of OFPMA to cause micronuclei in cultured human lymphocytes using cytochalasin B-block methodology with and without an exogenous metabolic activation system.

 

The test article was considered negative for inducing micronuclei in binucleated cells in cultured human lymphocytes without and with metabolic activation.

 

in vitro gene mutation study in mammalian cells

An in vitro gene mutation study in mammalian cells is the second part of the standard information set required for registration at the Annex VIII tonnage level.  ECHA’s Guidance on Information Requirements and Chemical Safety Assessment (ECHA, 2017) states it is possible to omit the second in vitro study in mammalian cells. If it can be demonstrated that this mammalian cell test will not provide any further useful information about the potential in vivo mutagenicity of a substance, then it does not need to be conducted.

Experimental data with OFPMA involving two in vitro gene mutation studies in bacteria (BioReliance Corporation, 2016; IIT Research Institute, 2008) and one in vitro micronucleus study (Covance Laboratories, 2009) are all negative; details are provided elsewhere within this dossier.   This demonstrates that OFPMA has no genotoxic potential and no further testing should be necessary.  This is consistent with the fact that the in vitro gene mutation study in bacteria (Ames assay) (OECD TG 471) and the in vitro micronucleus study (OECD TG 487) are the 2 in vitro genotoxicity tests recommended by the European Food Safety Authority for initial assessment of genotoxic potential of food additives or ingredients.  As stated in their guidance document (EFSA, 2012), should a compound be found to be without activity in the in vitro gene mutation study in bacteria and in the in vitro micronucleus assay, no further in vitro or in vivo testing is considered necessary.  The combination of the Ames and in vitro micronucleus tests has been demonstrated to be sufficient to detect almost all in vivo genotoxic and/or carcinogenic compounds and that addition of a gene mutation assay in mammalian cells (i.e., mouse lymphoma assay) adds little value in this regard (Kirkland et al., 2011).

An examination of the structure of the notified substance clearly shows that the substance comprises a short-chained polyfluoroalkyl methacrylate.  Assessment of the chemical structure of OFPMA has determined that the substance does not contain any structural alerts that are indicative of mutagenic, genotoxic or carcinogenic potential (Ashby and Tennant, 1991; Tennant and Ashby, 1991; Ashby, 1996; Benigni and Bossa, 2008).    

The substance is anticipated to be metabolized to octafluoropentanol and methacrylic acid.  The methacrylic acid is subsequently converted to carbon dioxide via the tricarboxylic acid cycle in both experimental animals and humans (WHO, 1998).  The octafluoropentanol is expected to be converted to octafluoropentanal via alcohol dehydrogenase and octafluoropentanoic acid via aldehyde dehydrogenase.  Assessment of the chemical structures of these metabolites likewise determined that the substances do not contain any structural alerts that are indicative of mutagenic, genotoxic or carcinogenic potential.

None of the chemical groups/moieties in the substance and its metabolites have been indicated as a structural alert for genotoxicity or for carcinogenicity.  Therefore, the in vitro mammalian cell test will not provide any further useful information about the potential in vivo mutagenicity of OFPMA.

REFERENCES

Ashby J (1996). Alternatives to the 2-species bioassay for the identification of potential human carcinogens. Human Exp Toxicol 15:183-202.

Ashby J, Tennant RW (1991). Definitive relationships among chemical structure, carcinogenicity and mutagenicity for 301 chemicals tested by the U.S. NTP (National Toxicology Program). Mutat Res 257(3):229-306.

Benigni R, Bossa C (2008). Structure alerts for carcinogenicity, and the Salmonella assay system: a novel insight through the chemical relational databases technology. Mutat Res 659(3):248-261

BioReliance Corporation (2016). Bacterial Reverse Mutation Assay. BioReliance Corporation, 9630 Medical Center Drive, Rockville, MD 20850. Unpublished report number AE46LA.503005.BTL (provided elsewhere within this dossier)

Covance Laboratories (2009). In Vitro Human Peripheral Blood Lymphocyte Micronucleus Assay. Covance Laboratories Inc., 9200 Leesburg Pike, Vienna, Virginia 22182-1699. Unpublished report number 8097-100. (provided elsewhere within this dossier)

ECHA (2017). Guidance on information requirements and chemical safety assessment. Chapter R.7a: Endpoint specific guidance.  Volume 3.0, July 2017.  Available at:  https://echa.europa.eu/documents/10162/13632/information_requirements_r7a_en.pdf/e4a2a18f-a2bd-4a04-ac6d-0ea425b2567f

EFSA (2012). Scientific Opinion. Scientific opinion on genotoxicity testing strategies applicable to food and feed safety assessment. EFSA Scientific Committee, European Food Safety Authority (EFSA), Parma, Italy.  EFSA Journal 9(9):2379.  Available at: http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2011.2379/epdf

IIT Research Institute (2008). Evaluation of the Potential Mutagenic Activity of OFPMA in the Bacterial Reverse Mutation Assay. IIT Research Institute, Life Sciences Group, 10 West 35th Street, Chicago, IL 60616-3799. Unpublished report number 2171-002. (provided elsewhere within this dossier)

Kirkland D, Reeve L, Gatehouse D, Vanparysd P (2011). A core in vitro genotoxicity battery comprising the Ames test plus the in vitro micronucleus test is sufficient to detect rodent carcinogens and in vivo genotoxins. Mutat Res 721(1):27-73

Tennant RW, Ashby J (1991). Classification according to chemical structure, mutagenicity to Salmonella and level of carcinogenicity of a further 39 chemicals tested for carcinogenicity by the U.S. National Toxicology Program. Mutat Res 257(3):209-227

WHO (1998). Concise International chemical assessment document 4. Methyl methacrylate. Geneva, Switzerland: World Health Organization (WHO). Available at: http://www.who.int/ipcs/publications/cicad/en/cicad04.pdf

Justification for classification or non-classification

The available in vitro studies demonstrate OFPMA is not mutagenic according to a bacterial reverse mutation assay and does not induce micronuclei in binucleated cells in cultured human lymphocytes without and with metabolic activation. Therefore, the experimental findings do not fulfil the criteria for germ cell mutagenicity classification according to European CLP Regulation (EC) No 1272/2008 (as amended).