Docosanoic acid, monoester with glycerol

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

From the literature search, specific data on mutagenicity of mono-glyceryl
monoesters were found in an assessment performed by the Cosmetic
Ingredient Review (CIR) expert panel (CIR 2016, 2019). Data on glycerol (CAS
No. 56-81-5) was identified from a European Food Safety Authorities (EFSA)
re-evaluation performed in 2017. Data from
Five in vitro mutagenic test were identified. Four out of the five studies are
Ames test, with and without metabolic activations, according to OECD
Guideline 471. Glyceryl laurate; glyceryl rosinate; acetic and fatty acid esters of
glycerol; glycerides, C14-18 and C16-22 unsaturated, mono- and di- were all
negative in the Ames test (positive controls gave expected results).
Undecylenic acid was tested for gene mutation both in vitro and in vivo test
systems. Undecylenic acid was not genotoxic in a Chinese hamster lung
fibroblast (V79), with and without metabolic activation; as well as not genotoxic
in male and female CD-1 mice. Glycerol tested negative for gene mutation
without metabolic activation in an HGPRT gene mutation assay performed on
Chinese hamster ovary cells.
One OECD 473 Guidelines test on chromosome aberration was identified for
glyceryl rosinate. Glycerol rosinate is a monoester of glycerol and long chain
fatty acids from rosin. Glyceryl rosinate was not genotoxic in the OECD 473
chromosome aberration test. Glyceryl acetate (89% pure) was also tested in an
in vitro gene mutation chromosomal aberration test (guideline study not stated).
Glyceryl acetate (89% pure) was not genotoxic in the tested system. Behenic
acid tested negative for genotoxicity in a mammalian chromosome aberrations
test at a concentration up to 3500 µg/mL; likewise at a similar concertation
Behenic acid (85.9% pure) tested negative for genotoxicity in an OECD 473
guideline study. Furthermore, glycerol tested negative for genotoxicity in three
independent in vitro mammalian chromosome aberration studies, and one in
vivo bone marrow chromosomal aberration test performed on male rats (strain
not specified).
Glycerides, C16-18 and C18-hydroxy mono- and di- were tested in an in vivo
micronucleus test, which gave negative results.
Glycerol was tested negative for DNA damage in an unscheduled DNA
synthesis assay and a sister chromatid exchange assay.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

other: weight of evidence analysis based on expert evaluated data on hydrolysis products and structural analogues

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Justification for type of information:

In relation to the data requirements of REACH Annex VIII (10-100 t/y), data on genotoxicity must be provided. Limited data on this endpoint is available for docosanoic acid, monoester with glycerol (glycerol monobehenate).

Glycerol monobehenate is a mono-constituent substance. The main component is docosanoic acid, monoester with glycerol which is present in the product at a concentration of 80 – 90%, the remaining compounds are mainly fatty acids and monoesters of fatty acid and glycerol. Glycerol can also be present in a low concentration. Glyceryl monoesters (monoglycerides) are metabolized to free fatty acids and glycerol, both of which are available for the resynthesis of triglycerides.

The mutagenicity of this substance is therefore assessed in the present document as a weight of evidence analysis based on existing data on groups of mono-, di- and triglycerides, fatty acids, which are all components with similar properties. Hereby, a huge data set is available for deriving a conclusion on the genotoxicity toxicity of the substance.

Principles of method if other than guideline:

In relation to the data requirements of REACH VIII (10-100 t/y), data on genetic toxicity must be provided. Limited data on this endpoint is available for Docosanoic acid, monoester with glycerol (glycerol monobehenate). Glycerol monobehenate is a mono-constituent substance. The main component is docosanoic acid, monoester with glycerol, the remaining compounds are mainly fatty acids and monoesters of fatty acid and glycerol. Glycerol can also be present in a low concentration. Glyceryl monoesters (monoglycerides) are metabolized to free fatty acids and glycerol, both of which are available for the resynthesis of triglycerides.

The possible genetic toxicity of this substance is therefore assessed in the present weight of evidence analysis based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, and The Danish EPA QSAR database), and data generated from similar chemicals

Expert group assessments of the substances are considered the most valid data for the assessment, an overall weight of evidence approach based on these expert evaluations is used for the assessment.

Type of assay:

in vitro mammalian chromosome aberration test

Key result

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Conclusions:

Limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). However, based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish EPA QSAR database) and data generated from similar chemicals it is concluded that the Docosanoic acid, monoester with glycerol (glycerol monobehenate) is considered not be a clastogenic substance as no chromosome aberrations was seen in an OECD 473 study.

Executive summary:

From the literature search, specific data on mutagenicity of mono-glyceryl monoesters were found in an assessment performed by the Cosmetic Ingredient Review (CIR) expert panel (CIR 2016, 2019). 

Undecylenic acid was tested for gene mutation both in vitro and in vivo test systems. Undecylenic acid was not genotoxic in a Chinese hamster lung fibroblast (V79), with and without metabolic activation; as well as not genotoxic in male and female CD-1 mice. Glycerol tested negative for gene mutation without metabolic activation in an HGPRT gene mutation assay performed on Chinese hamster ovary cells. One OECD 473 Guidelines test on chromosome aberration was identified for glyceryl rosinate. Glycerol rosinate is a monoester of glycerol and long chain fatty acids from rosin. Glyceryl rosinate was not genotoxic in the OECD 473 chromosome aberration test. Glyceryl acetate (89% pure) was also tested in an in vitro gene mutation chromosomal aberration test (guideline study not stated). Glyceryl acetate (89% pure) was not genotoxic in the tested system. Behenic acid tested negative for genotoxicity in a mammalian chromosome aberrations test at a concentration up to 3500 µg/mL; likewise at a similar concertation Behenic acid (85.9% pure) tested negative for genotoxicity in an OECD 473 guideline study. Furthermore, glycerol tested negative for genotoxicity in three independent in vitro mammalian chromosome aberration studies, and one in vivo bone marrow chromosomal aberration test performed on male rats (strain not specified). Glycerides, C16-18 and C18-hydroxy mono- and di- were tested in an in vivo micronucleus test, which gave negative results. Glycerol was tested negative for DNA damage in an unscheduled DNA synthesis assay and a sister chromatid exchange assay.

Further, EFSA 2020 concluded that glycerol monobehenate was found to be non-genotoxic based on QSAR predictions using the OECD QSAR ToolBox and QSAR VEGA Tool.

In summary, limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). However, based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish EPA QSAR database) and data generated from similar chemicals it is concluded that the Docosanoic acid, monoester with glycerol (glycerol monobehenate) is considered not be genotoxic and not a clastogenic substance as no chromosome aberrations was seen in vitro and in vivo. 

Reference 2

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

other: weight of evidence analysis based on expert evaluated data on hydrolysis products and structural analogues

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Justification for type of information:

Glycerol monobehenate is a mono-constituent substance. The main component is docosanoic acid, monoester with glycerol, the remaining compounds are mainly fatty acids and monoesters of fatty acid and glycerol. Glycerol can also be present in a low concentration. Glyceryl monoesters (monoglycerides) are metabolized to free fatty acids and glycerol, both of which are available for the resynthesis of triglycerides.

Limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). Therefore, the genotoxicity assessment of the substance was based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, and The Danish EPA QSAR database), and data generated from similar chemicals

A weight of evidence document is attached. The conclusion in this document is based on data from the following expert assessments:

- CIR 2016: Cosmetic Ingredient Review. Safety assessment of monoglyceryl monoesters as used in cosmetics. Final amended report, January 15, 2016.  
- EFSA 2020: Review and priority setting for substances that are listed without a specific migration limit in Table 1 of Annex 1 of Regulation 10/2011 on plastic materials and articles intended to come into contact with food. EFSA Journal 2020;18(6):6124.
- BIAC/ICCA, CoCAM 6, 30 – 03 October 2014. SIDS Initial assessment profile.

Principles of method if other than guideline:

In relation to the data requirements of REACH Annex VIII (10-100 t/y), data on genotoxicity must be provided. Limited data on this endpoint is available for docosanoic acid, monoester with glycerol (glycerol monobehenate).

Glycerol monobehenate is a mono-constituent substance. The main component is docosanoic acid, monoester with glycerol which is present in the product at a concentration of 80 – 90%, the remaining compounds are mainly fatty acids and monoesters of fatty acid and glycerol. Glycerol can also be present in a low concentration. Glyceryl monoesters (monoglycerides) are metabolized to free fatty acids and glycerol, both of which are available for the resynthesis of triglycerides.

The mutagenicity of this substance is therefore assessed in the present document as a weight of evidence analysis based on existing data on groups of mono-, di- and triglycerides, fatty acids, which are all components with similar properties. Hereby, a huge data set is available for deriving a conclusion on the genotoxicity toxicity of the substance.

Type of assay:

other: Mammalian gene mutation assay, with and without metabolic activation, Chinese hamster lung fibroblast (V79)

Key result

Species / strain:

Chinese hamster lung (CHL/IU)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Conclusions:

Limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). However, based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish EPA QSAR database) and data generated from similar chemicals it is concluded that the Docosanoic acid, monoester with glycerol (glycerol monobehenate) is considered not genotoxic.

Executive summary:

From the literature search, specific data on mutagenicity of mono-glyceryl monoesters were found in an assessment performed by the Cosmetic Ingredient Review (CIR) expert panel (CIR 2016, 2019). 

Undecylenic acid was tested for gene mutation both in vitro and in vivo test systems. Undecylenic acid was not genotoxic in a Chinese hamster lung fibroblast (V79), with and without metabolic activation; as well as not genotoxic in male and female CD-1 mice. Glycerol tested negative for gene mutation without metabolic activation in an HGPRT gene mutation assay performed on Chinese hamster ovary cells. One OECD 473 Guidelines test on chromosome aberration was identified for glyceryl rosinate. Glycerol rosinate is a monoester of glycerol and long chain fatty acids from rosin. Glyceryl rosinate was not genotoxic in the OECD 473 chromosome aberration test. Glyceryl acetate (89% pure) was also tested in an in vitro gene mutation chromosomal aberration test (guideline study not stated). Glyceryl acetate (89% pure) was not genotoxic in the tested system. Behenic acid tested negative for genotoxicity in a mammalian chromosome aberrations test at a concentration up to 3500 µg/mL; likewise at a similar concertation Behenic acid (85.9% pure) tested negative for genotoxicity in an OECD 473 guideline study. Furthermore, glycerol tested negative for genotoxicity in three independent in vitro mammalian chromosome aberration studies, and one in vivo bone marrow chromosomal aberration test performed on male rats (strain not specified). Glycerides, C16-18 and C18-hydroxy mono- and di- were tested in an in vivo micronucleus test, which gave negative results. Glycerol was tested negative for DNA damage in an unscheduled DNA synthesis assay and a sister chromatid exchange assay.

Further, EFSA 2020 concluded that glycerol monobehenate was found to be non-genotoxic based on QSAR predictions using the OECD QSAR ToolBox and QSAR VEGA Tool.

 

In summary, limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). However, based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish EPA QSAR database) and data generated from similar chemicals it is concluded that the Docosanoic acid, monoester with glycerol (glycerol monobehenate) is considered not be genotoxic in vitro. 

Reference 3

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

other: Weight of evidence analysis based on expert reviews on hydrolysis products and structural analogues

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: based on expert group reviews

Justification for type of information:

Glycerol monobehenate is a mono-constituent substance. The main component is docosanoic acid, monoester with glycerol, the remaining compounds are mainly fatty acids and monoesters of fatty acid and glycerol. Glycerol can also be present in a low concentration. Glyceryl monoesters (monoglycerides) are metabolized to free fatty acids and glycerol, both of which are available for the resynthesis of triglycerides.

Limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). Therefore, the genotoxicity assessment of the substance was based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, and The Danish EPA QSAR database), and data generated from similar chemicals

A weight of evidence document is attached. The conclusion in this document is based on data from the following expert assessments:

- CIR 2016: Cosmetic Ingredient Review. Safety assessment of monoglyceryl monoesters as used in cosmetics. Final amended report, January 15, 2016.  
- EFSA 2020: Review and priority setting for substances that are listed without a specific migration limit in Table 1 of Annex 1 of Regulation 10/2011 on plastic materials and articles intended to come into contact with food. EFSA Journal 2020;18(6):6124.
- BIAC/ICCA, CoCAM 6, 30 – 03 October 2014. SIDS Initial assessment profile.

Principles of method if other than guideline:

In relation to the data requirements of REACH VIII (1-10 t/y), data on genetic toxicity must be provided. Limited data on this endpoint is available for Docosanoic acid, monoester with glycerol (glycerol monobehenate). Glycerol monobehenate is a mono-constituent substance. The main component is docosanoic acid, monoester with glycerol, the remaining compounds are mainly fatty acids and monoesters of fatty acid and glycerol. Glycerol can also be present in a low concentration. Glyceryl monoesters (monoglycerides) are metabolized to free fatty acids and glycerol, both of which are available for the resynthesis of triglycerides.

The possible genetic toxicity of this substance is therefore assessed in the present weight of evidence analysis based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, and The Danish EPA QSAR database), and data generated from similar chemicals

Expert group assessments of the substances are considered the most valid data for the assessment, an overall weight of evidence approach based on these expert evaluations is used for the assessment.

Type of assay:

other: weight of evidence analysis based on expert evaluated data, prediction data and dat from structural analogues

Key result

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Key result

Species / strain:

other: QSAR prediction

Metabolic activation:

not applicable

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

other: Not applicable

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:

Further, five in vitro mutagenic test were identified. Four out of the five studies are Ames test, with and without metabolic activations, according to OECD Guideline 471. Glyceryl laurate; glyceryl rosinate; acetic and fatty acid esters of glycerol; glycerides, C14-18 and C16-22 unsaturated, mono- and di- were all negative in the Ames test (positive controls gave expected results).

The attached weight of evidence document describes in silico prediction data for Docosanoic acid, monoester with glycerol (glycerol monobehenate) using a number of QSAR models as well as Ames testing data for a number of similar monoglyceryl monoesters and mono-, di- and triglycerides. Please see the attached document for details.

Data from QSAR models are described below:

- Danish Q(SAR) database (accessed 6-1-2021): An assessment report of Docosanoic acid, monoester with glycerol (glycerol monobehenate) was identified in the Danish EPA Q(SAR) database. No structural alerts for DNA reactivity were found.Furthermore, the main Ames model (Ames test in S. Typhimurium(in vitro)) was negative, all other genotoxicity in vitro test also showed negative results for genotoxicity.

- EFSA 2020: In a review for substances that are listed without a specific migration limit in Table 1 of Annex 1 of Regulation 10/2011 on plastic materials and articles intended to come into contact with food, EFSA concluded that glycerol monobehenate was found to be non-genotoxic based on QSAR predictions using the OECD QSAR ToolBox and QSAR VEGA Tool. The predictions were considered valid and inside the applicability domain. Specifically, the results from the OECD QSAR Toolbox showed no alert for In vitro mutagenicity (Ames test) alerts by ISS and no alert for DNA alerts for AMES by OASIS. Further, specifically the results using the QSAR VEGA Tool were Consensus model: Non-Mutagenic and Non-mutagenic score 0.675.

Data from bacterial mutation assay (Ames) are described below:

- No testing data was identified for docosanoic acid, monoester with glycerol (glycerol monobehenate). Ames testing data from similar monoglyceryl monoesters (Glyceryl acetate, Glyceryl laurate, Glyceryl rosinate) all points towards no genotoxicity in vitro (CIR 2016).

- Further, Ames testing data from mono, di- and triglycerides also points towards no genotoxicity in vitro (OECD SIDS 2014).

 

Conclusions:

Limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). However, based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish EPA QSAR database) and testing data available from comparable monoglyceryl monoesters as well as mono. di- and tri-glycerides, it is concluded that Docosanoic acid, monoester with glycerol (glycerol monobehenate) is considered non genotoxic in vitro in relation to the bacterial reverse mutation test (Ames).

Executive summary:

No testing data was identified for docosanoic acid, monoester with glycerol (glycerol monobehenate). Ames testing data from similar monoglyceryl monoesters (Glyceryl acetate, Glyceryl laurate, Glyceryl rosinate) all points towards no genotoxicity in vitro (CIR 2016). Further, Ames testing data from mono, di- and triglycerides also points towards no genotoxicity in vitro (OECD SIDS 2014).

Further, from the literature search, specific data on mutagenicity of mono-glyceryl monoesters were found in an assessment performed by the Cosmetic Ingredient Review (CIR) expert panel (CIR 2016, 2019). Data on glycerol (CAS No. 56-81-5) was identified from a European Food Safety Authorities (EFSA) re-evaluation performed in 2017.

 

Five in vitro mutagenic test were identified. Four out of the five studies are Ames test, with and without metabolic activations, according to OECD Guideline 471. Glyceryl laurate; glyceryl rosinate; acetic and fatty acid esters of glycerol; glycerides, C14-18 and C16-22 unsaturated, mono- and di- were all negative in the Ames test (positive controls gave expected results).

In support, the possible genetic toxicity in vitro of glycerol monobehenatewas also  assessed in the present weight of evidence analysis based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, and The Danish EPA QSAR database), and data generated from similar chemicals. No structural alerts for DNA reactivity were found. Furthermore, the main Ames model (Ames test in S.
Typhimurium(in vitro)) was negative,

 

 

In conclusion, limited in vitro and in vivo genotoxicity data was identified for Docosanoic acid, monoester with glycerol (glycerol monobehenate). However, based on Q(SAR) predictions using different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish EPA QSAR database) and data generated from similar chemicals it is concluded that the Docosanoic acid, monoester with glycerol (glycerol monobehenate) is considered not genotoxic in vitro in relation to bacterial reverse mutation (Ames test).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Sufficient test data has been identified for mutagenicity. It is concluded, based
on this weight of evidence, that there are no signs of genotoxic effect from the
impurities (i.e., Glycerol (56-81-5); Stearic acid; Docosanoic acid; Glycerol
mono esters with C14, C16, C18, C20 and C24 saturated fatty acids) as well
as from other similar substances as Docosanoic acid, monoester with glycerol
(glycerol monobehenate). Furthermore, based on Q(SAR) predictions using
different QSAR models (OECD QSAR Toolbox, VEGA QSAR Tool, The Danish
EPA QSAR database) and data generated from similar chemicals it is
concluded that the Docosanoic acid, monoester with glycerol (glycerol
monobehenate) is considered not genotoxic.

 

 

Thus,the overall weight of evidence indicates that glycerol monobehenate does not cause genotoxicity in vitro and therefore, the substance should not be classified for this endpoint according to (EC) No 1272/2008.