Isooctadecanoic acid, monoester with glycerol

Endpoint summary

Additional information

Analogue justification

Data on the genetic toxicity of Glycerol monoisostearate (CAS 66085-00-5) are not available. The assessment was therefore based on studies conducted with analogue substances as part of a read across approach, which is in accordance with Regulation (EC) No. 1907/2006, Annex XI, 1.5. For each specific endpoint the source substance(s) structurally closest to the target substance is/are chosen for read-across, with due regard to the requirements of adequacy and reliability of the available data. Structural similarities and similarities in properties and/or activities of the source and target substance are the basis of read-across. A detailed justification for the grouping of chemicals and read-across is provided in the technical dossier (see IUCLID Section 13) and within Chapter 5.1 of the CSR.

Genetic toxicity (mutagenicity) in bacteria in vitro

CAS No. 111-03-5

In a GLP-study performed according to OECD guideline 471, the potential mutagenicity of 2,3-dihydroxypropyl oleate was investigated in S. typhimurium TA 1535, TA 1537, TA 98 and TA 100 and in E. coli WP2 uvrA (Nakajima, 2005). In three dose finding tests with test concentrations up to 5000 µg/plate, suitable concentrations for treatment of the bacterial strains in the main study were selected. Based on the results from pre-test on solubility and/or cytotoxicity of the test substance, concentrations up to 9.77 (TA 1537), 156 (TA 98 and TA 100) and 5000 µg/plate (TA 1535, WP2 uvrA) were chosen in the absence of metabolic activation, while concentrations up to 313 (TA 1537), 625 (TA 100) and 5000 µg/plate (TA 1535, WP2 uvrA and TA 98) were chosen in the presence of metabolic activation, respectively. In the main experiment, cytotoxic effects were observed at 4.88 (TA 1537), 78.1 (TA 98) and 156 (TA 100) µg/plate without metabolic activation, as well as at 156 (TA 100) and 313 (TA 1537) µg/plate with metabolic activation. Precipitation was noted at the end of the exposure period at ≥ 625 and ≥ 1250 µg/plate in the presence and absence of metabolic activation, respectively. No increase in the mean number of revertants per plate was observed in any strain compared with the controls. The positive and negative controls included in the assay demonstrated the validity of the assay. Under the conditions of this assay, the test substance was not mutagenic in the presence or absence of metabolic activation.

CAS No. 26942-95-0

The potential mutagenicity of 1,2,3-propanetriyl triisooctadecanoate was assessed in four S. typhimurium strains (TA 1535, TA 1537, TA 98 and TA 100) in an Ames test similar to OECD Guideline 471 and under GLP conditions (Waart van de, 1996). Concentrations ranging from 100 to 5000 µg/plate were selected for treatment in the main assay. Precipitation was observed in the first experiment at the maximum concentration of 5000 µg/plate, which however did not influence the counting of revertant colonies. No increase in the mean number of revertants per plate was observed when compared to controls. The positive and negative controls included for each tester strain were shown to be valid. Based on the study results, the test substance was considered non-mutagenic in the selected strains of S. typhimurium in the presence and absence of metabolic activation.

CAS No. 91845-19-1

A bacterial gene mutation assay with Glycerides, C16-18 and C18-hydroxy mono- and di- was performed in compliance with OECD guideline 471 and GLP (Wallat, 1984). In two independent experiments, S. typhimurium TA 1535, TA 1537, TA 1538, TA 98 and TA 100 were exposed to concentrations from 8 to 5000 µg/plate with and without metabolic activation. At the maximum concentration, bacterial growth was completely inhibited in Salmonella strain TA 1537 in the presence (first experiment) or absence (second experiment) of metabolic activation, respectively. No increase in the mean number of revertants per plate was observed in any tester strain at the indicated concentrations. The vehicle controls were within the spontaneous mutation ranges of the historical controls and the positive controls induced the expected increase in the number of reverse mutants. Under the conditions of this experiment, the test substance did not show mutagenicity in the selected strains of S. typhimurium in the presence and absence of metabolic activation.

EC No. 460-300-6

A bacterial gene mutation assay with Glycerides, C16-18 and C18-unsaturated, branched and linear mono-, di- and tri- was performed in compliance with OECD guideline 471 and GLP (Bowles, 2004). In a preliminary cytotoxicity test, no cytotoxicity was observed in TA 100 at up to 5000 µg/plate with and without metabolic activation system (S9-mix). Thus, concentrations up to 5000 µg/plate were applied for treatment of S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102 in two independent experiments in the presence or absence of metabolic activation. An oily precipitate was observed at 5000 µg/plate (with and without S9-mix) in all experiments, but this did not interfere with the scoring of revertant colonies. The mean number of revertant colonies per plate was not increased in any tester strain and at any test concentration with or without metabolic activation. The mean numbers of spontaneous revertants in the negative (solvent) controls and the mean number of revertants in the positive controls of the strains used were all within the normal historical ranges. Therefore, the test substance was considered to be non-mutagenic in the selected strains of S. typhimurium under the conditions of this Ames test.

Genetic toxicity (cytogenicity) in mammalian cells in vitro

CAS No. 111-03-5

2,3-Dihydroxypropyl oleate was assessed in an in vitro mammalian chromosome aberration test in Chinese hamster lung cells (CHL/IU) according to OECD guideline 473 and under GLP conditions (Nakajima, 2005). In a preliminary cytotoxicity test, cells were exposed to concentrations ranging from 6.96 to 3565 µg/mL for a continuous 24-h exposure period with metabolic activation (S9-mix) or a short-term 6 h exposure with and without S9-mix. The test substance was not cytotoxic after short-term exposure at any concentration in the presence of S9 mix. Without metabolic activation, relative growth was reduced to about 71% of the control value after a period of 6 h. Continuous exposure for 24 h without S9-mix caused no cytotoxicity up to concentrations of 1783 µg/mL. At 3565 µg/mL, relative growth was decreased to ca. 13% of the control value. The concentration leading to 50% cytotoxicity was calculated to be 2738 µg/mL. Based on these results, concentrations of 891, 1783 and 3565 µg/mL (with and without S9 mix) were selected for chromosome analysis after short-term exposure, whereas concentrations of 55.7, 111, 223 and 446 µg/mL (without S9-mix) were chosen for chromosome analysis after continuous exposure. No increase in the number of cells with chromosomal aberrations was observed compared to controls in any of the experiments performed. No cytotoxic effects were observed after short-term exposure, but the test substance was cytotoxic at 446 µg/mL after 24-h continuous treatment. Visible precipitation of the test substance was observed at concentrations ≥ 223 µg/mL, however, this did not interfere with chromosomal analysis. The positive controls included during short-term and continuous exposure were shown to be valid. Under the conditions of this experiment, the test substance was considered to be not clastogenic in Chinese hamster lung cells (CHL/IU) in the presence and absence of metabolic activation.

CAS No. 736150-63-3

An in vitro mammalian chromosome aberration test was performed with Glycerides, castor-oil.mono, hydrogenated, acetates in human lymphocytes according to OECD guideline 473 and in compliance with GLP (Edwards, 2004). An initial cytotoxicity test was performed with concentrations of 313 - 5000 µg/mL in the presence or absence of metabolic activation (S9-mix) for 3 h. A dose-related toxicity was observed. At the highest concentration (5000 µg/mL), the mitotic index was reduced to 64% of the vehicle control without metabolic activation and to 10% with metabolic activation, respectively. Based on these results, concentrations of 625, 1250, 2500 and 5000 µg/mL (without S9-mix) and 625, 1250, 2500, 3600 and 5000 µg/mL (with 2% S9-mix) were chosen for treatment of cells in the main assay for an exposure period of 3 h. Metaphase analysis was performed at concentrations of 1250, 2500 and 5000 µg/mL (without S9-mix) and 625, 1250 and 2500 µg/mL (with 2% S9-mix). In the second main assay, cells were exposed to concentrations of 313-5000 µg/mL (without S9 mix) or 625-5000 µg/mL (with 4% S9 mix) for either 3 or 20 h, respectively. As the samples without metabolic activation revealed mean mitotic indices lower than 50% of the solvent control in all dose groups, this part of the experiment was repeated with lower concentrations ranging from 2.5 to 320 µg/mL. Based on the cytotoxicity data obtained, concentrations of 40, 80 and 160 µg/mL (without S9) and 625, 1250 and 2500 µg/mL (with 4% S9-mix) were used for metaphase analysis. The chromosome analysis of the experiments showed no treatment-related increase in the number of cells with chromosomal aberrations compared with controls. The frequency of metaphases with chromosomal aberrations in the solvent controls was compatible to the historical control values and the positive controls were shown to be valid. Based on the results of this chromosome aberration test, the test substance was not clastogenic in human lymphocytes in the presence or absence of metabolic activation under the experimental conditions chosen.

Short-, medium- and long-chain triglycerides (SCT, MCT, LCT)

An in vitro mammalian chromosome aberration test with the SALATRIM (short- and long-chain acyl triglyceride molecules) family of triacylglycerols was performed similar to OECD guideline 476 in Chinese hamster Ovary (CHO) cells (Hayes et al., 1994). In a preliminary toxicity test, concentrations of 8 - 1000 μg/mL were used to determine suitable concentrations for chromosome analysis. Based on these results, the occurrence of chromosome aberrations was investigated in the presence and absence of metabolic activation (rat liver S9-mix) with test substance concentrations of 250, 500 and 1000 µg/mL. The exposure duration was 2 and 8 hours with and without metabolic activation, respectively, while the harvest time was 8-10 h (with and without metabolic activation). The highest dose was chosen based on the low solubility of the fats in the assay medium. No significant increase in the number of phases with aberrations was observed in treated CHO cells at any preparation time and concentration. No significant cytotoxic effects were reported. The positive controls significantly increased the rate of chromosome aberrations. In conclusion, the test substance was not clastogenic in Chinese hamster ovary cells, neither in the presence nor in the absence of a metabolic activation system, under the experimental conditions chosen.

Genetic toxicity (mutagenicity) in mammalian cells in vitro

CAS No. 736150-63-3

An in vitro mammalian cell gene mutation assay was performed with Glycerides, castor-oil-mono, hydrogenated, acetates according to OECD guideline 476 and under GLP conditions (Edwards, 2002). In the first experiment, mutations at the TK locus of mouse-lymphoma L5178Y cells were investigated at concentrations of 625, 1250, 2500, 3600 and 5000 µg/mL. The L5178Y cells were exposed to the test material for a period of 3 h in the presence and for 4 h in the absence of metabolic activation (S9-mix), respectively. At 3600 µg/mL, the relative total growth was 1-11% compared with the negative controls. In the second experiment, cells were exposed to a concentration range of 313 to 3600 µg/ for a period of 24 h, without S9-mix, and to a concentration range of 156-3600 µg/mL for a period of 4 h, with S9-mix. Since the relative growth without S9-mix was very low (0-2%) at all test concentrations, the 24-h treatment of cells in the absence of S9-mix was repeated with concentrations ranging from 2.5-320 µg/mL, which resulted in appropriate levels of cytotoxicity (10-20% relative growth) at 160 µg/mL. In the presence of metabolic activation, the relative total growth was 37 and 0% at 2500 and 3600 µg/mL in the second experiment, respectively. After a 3-day expression period of the cultures, the resistance to 5-trifluorothymidine (TFT) was determined in all experiments. The test substance did not induce a significant increase in the mutant frequency at any preparation time and dose concentration. The positive controls significantly increased mutant frequency. In conclusion, the test substance did not induce mutations in mouse-lymphoma L5178Y cells, neither in the presence nor in the absence of a metabolic activation system, under these experimental conditions.

Short-, medium- and long-chain triglycerides (SCT, MCT, LCT)

The SALATRIM (short- and long-chain acyl triglyceride molecules) family of triacylglycerols was assessed using a gene mutation assay in cultured mammalian cells (Chinese hamster ovary (CHO-K1) cells) similarly to OECD guideline 476 (Hayes et al., 1994). Gene mutations at the HPRT locus were investigated in the presence and absence of metabolic activation (rat liver S9-mix) with concentrations of 31.25, 62.5, 125, 250, 500 and 1000 µg/mL. Two cultures were tested per dose level. The highest concentration was limited by the low solubility of the fats in the assay medium. No significant cytotoxicity was reported. An increase in mutant frequency was not observed at any concentration tested whether with or without metabolic activation. The positive controls significantly increased the mutant frequency. Therefore, it was concluded that under the conditions of the study, the test material was not mutagenic at the HPRT locus of Chinese hamster ovary cells in the absence and presence of metabolic activation.

Overall conclusion for genetic toxicity

There are no available studies on the genetic toxicity of Glycerol monoisostearate. Therefore analogue read-across from source substances was applied.

The available data on the genetic toxicity of 6 source substances includes several studies on the potential induction of gene mutations in bacteria as well as several in vitro studies on cytogenicity and gene mutations in mammalian cells. The results of the available in vitro studies were negative.

Based on the available data and following the analogue approach, Glycerol monoisostearate is considered to be not mutagenic in vitro and not clastogenic in vitro.

Justification for selection of genetic toxicity endpoint
No study was selected, as all the in vitro studies were negative.

Short description of key information:
No mutagenicity in bacteria was observed in the available studies.
No clastogenic effects in mammalian cells were observed in the available studies.
No mutagenicity in mammalian cells was observed in the available studies.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

According to Article 13 of Regulation (EC) No. 1907/2006 "General Requirements for Generation of Information on Intrinsic Properties of substances", information on intrinsic properties of substances may be generated by means other than tests e.g. from information from structurally related substances (grouping or read-across), provided that conditions set out in Annex XI are met. Annex XI, "General rules for adaptation of this standard testing regime set out in Annexes VII to X” states that “substances whose physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity may be considered as a group, or ‘category’ of substances. This avoids the need to test every substance for every endpoint". Since the analogue concept is applied to Glycerol monoisostearate (CAS No. 66085-00-5), data will be generated from data for reference source substance(s) to avoid unnecessary animal testing. Additionally, once the analogue read-across concept is applied, substances will be classified and labelled on this basis.

Therefore, based on the analogue read-across approach, the available data on genetic toxicity do not meet the classification criteria according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.