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Long-term toxicity to fish

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Description of key information

The chemical safety assessment according to Annex I of Regulation (EC) No. 1907/2006 does not indicate the need to investigate further the long-term toxicity to fish.

Key value for chemical safety assessment

Additional information

No experimental data evaluating the chronic toxicity of Glycerides, C12-18 mono- and di- (CAS No. 91052-49-2) to fish species are available. According to Regulation (EC) No. 1907/2006, Annex IX, 9.1.5, long-term toxicity testing shall be proposed by the registrant if the chemical safety assessment according to Annex I indicates the need to investigate further the effects on aquatic organisms. This is not the case for Glycerides, C12-18 mono- and di-, as all available information indicates no concern related to long-term toxicity, as discussed below.


Glycerides, C12-18 mono- and di- is readily biodegradable (79.5% biodegradation in 28 days) and it has potential for adsorption (log Koc values 1.46-7.96). According to the Guidance on information requirements and chemical safety assessment, Chapter R.7b, readily biodegradable substances can be expected to undergo rapid and ultimate degradation in most environments, including biological Sewage Treatment Plants (STPs)(ECHA, 2012). Besides being extensively biodegraded in STPs (due to its ready biodegradability), a significant degree of removal of this substance from the water column due to adsorption to sewage sludge can be expected (Guidance on information requirements and chemical safety assessment, Chapter R.7a (ECHA, 2012)). Therefore, after passing through conventional STPs, only low concentrations of these substances are likely to be (if at all) released into the environment.


Moreover, rapid metabolization of Glycerides, C12-18 mono- and di-in aquatic organisms is expected. Enzymatic hydrolysis is expected to result in C12-18 fatty acids and glycerol as transformation products. Part of the free fatty acids will be re-sterified with glycerol and partial acyl glycerols to form triglycerides that will be stored as long-term energy reserves (Tocher, 2003). Glycerol is naturally present in animal and vegetable fats, rarely found in free state (mostly combined with fatty acids forming triglycerides) (ed. Knothe, van Gerpen and Krahl, 2005). If freely available in aquatic organisms, it will not bioaccumulate in view of its log Kow value of -1.76 (OECD SIDS, 2002). Especially in periods in which the energy demand is high (reproduction, migration, etc.), glycerides are mobilized from the storage sites as source of fatty acids. Fatty acid catabolism is the most important energy source in many species of fish, resulting in the release of acetyl CoA and NADH (through β-oxidation) and eventually, via the tricarboxylic cycle, the production of metabolic energy in the form of ATP. This fatty acid-catabolism pathway is the predominant source of energy related to growth, reproduction and development from egg to adult fish. A similar metabolic pathway is observed in mammals (see section 7.1.1 Basic toxicokinetics). According to the Guidance on information requirements and chemical safety assessment, Chapter R.7c (ECHA, 2012), even though ready biodegradability does not per se preclude bioaccumulation potential, generally (depending on exposure and uptake rates) ready biodegradable substances are likely to be rapidly metabolised, and therefore, concentrations stored in aquatic organisms will tend to be low. Considering the above information, low bioaccumulation potential of Glycerides, C12-18 mono- and di- in aquatic organisms can be expected.


The acute toxicity study conducted with Glycerides, C12-18 mono- and di- on fish showed no effects up to the water solubility of the substance, with a LL50 > 20 mg/L (loading rate), corresponding to a measured concentration of 2.7 mg/L. Effects occurred above the water solubility of the substance in the tests conducted with aquatic invertebrates and algae. The EL50 (48 h) value in the test performed with Daphnia magna was determined to be 36.2 mg/L (loading rate), corresponding to a measured concentration of 5.6 mg/L. The test performed with Desmodesmus subspicatus resulted in an EL50 (72 h) of 13 mg/L (based on growth rate, loading rate) and a NOELR (72 h) of 4.1 mg/L (based on growth rate, loading rate). Nevertheless, the possibility of physical effects in these two tests (due to disturbance of test organisms with emulsified test substance) instead of, or in addition to toxicological effects, cannot be excluded. The test material was reported to be sparingly soluble and therefore Water Accomodated Fractions (WAFs) were prepared in the algae and aquatic invertebrates tests. The WAFs were stirred for 48 hours and thereafter left for sedimentation for a period of 1 hour. After the sedimentation period, all WAFs in which effects were observed later on, were reported to be turbid. The WAFs were not filtered for the final tests. The highest loading rates in the algae test (45.7 mg/L and 99.4 mg/L) contained smaller algae cells, which had a different shape (crumpled) compared to those in the control. Furthermore, at these two loading rates, a dense emulsion of oily drops was observed.


Scientific evidence showed that aquatic toxicity testing of this type of Glycerides is technically very difficult. In an article by Prajapati et al. (2012)(see IUCLID section 6.1.4), the phase behaviour of lipid/surfactant/water phases was investigated, where medium-chain (C8-10) mono-, di- and triglycerides represent the lipid. Phase boundaries between lipids (monoglycerides, diglycerides, triglycerides), surfactant (PEG-35 castor oil) and water were established by visual inspection after an equilibration period, and the results expressed in phase diagrams. Viscosity and particle size distribution were measured. The mixtures with monoglyceride displayed two predominant phases: microemulsion and emulsion phases, whereas di- and triglycerides showed additionally a gel phase. Mixtures of monoglycerides and diglycerides, and of monoglycerides and triglycerides seemed to promote an increase of the microemulsion phase (in the 4 phases equilibrium). Particle size in these mixtures was found to be much smaller than in the monoglyceride sample alone. Microemulsions are solutions with an average particle size < 0.2 µm. This particle size would not be intercepted by a standard filter used in an aquatic toxicity test (generally, pore size of 0.45 µm). Due to their small size, based on visual inspection, clear or translucent solutions might be observed even when these microemulsions are present. Glycerides, C12-18 mono- and di- contains 40-70% C12 fatty acids and formation of microemulsions in test solutions is therefore possible for this substance. The WAFs at which effects were observed in the acute tests conducted with this substance (aquatic invertebrates and algae) were reported to be turbid, even though suspended particles could not be directly observed in the aquatic invertebrates test. This is considered as an indication of a microemulsion-formation phase, showing that the observed effects might indeed be due to physical interference with emulsified test material. The chances of microemulsion formation under chronic exposure to this substance are very high. Due to the prolonged testing period for these long-term toxicity tests, microemulsions can cause physical effects on fish (e.g gill clogging) and aquatic invertebrates (e.g. physical entrapment). Therefore, obtaining reliable chronic values out of such tests is technically difficult. Furthermore, based on the results of the acute tests, there is no indication showing that Glycerides, C12-18 mono- and di- would be toxic to fish, aquatic invertebrates and algae up to the limit of its water solubility. Nevertheless, a NOELR value is available for the species showing the highest sensitivity (algae) and is considered for hazard assessment as a worst-case approach (toxicity cannot be completely ruled out). Since the nominal NOELR value of 4.1 mg/L is just above the water solubility of the substance (measured concentration was below the LOD), the water solubility value of Glycerides, C12-18 mono- and di- (3.3 mg/L) will be taken for PNEC derivation, as a worst-case value.


Considering the expected low bioavailability of the substance in water, its low bioaccumulation potential, the technical difficulties involved in aquatic toxicity testing of medium-chain Glycerides (due to their tendency to form microemulsions), and the fact that a NOELR value is available for the species showing the highest sensitivity (algae), long-term toxicity testing is not deemed necessary for Glycerides, C12-18 mono- and di- (CAS No. 91052-49-2).


A detailed reference list is provided in the technical dossier (see IUCLID, section 13) and within the CSR.