2-ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate

Endpoint summary

Additional information

Basic toxicokinetics

There are no studies available in which the toxicokinetic behaviour of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate (CAS No. 25111-05-1) has been investigated.

Therefore, in accordance with Annex VIII, Column 1, Item 8.8 of Regulation (EC) 1907/2006 and with Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance (ECHA, 2012), assessment of the toxicokinetic behaviour of the substance 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate was conducted to the extent that can be derived from the relevant available information on physico-chemical and toxicological characteristics of structural analogue substances.

This comprises a qualitative assessment of the available substance-specific data on physico-chemical and toxicological properties according to Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance‘ (ECHA, 2012) and taking into account further available information on structural analogue substances from which data was used for read-across to cover data gaps.

The substance 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is an organic liquid composed of esters of trimethylolpropane (TMP) and mainly C18:1 unsaturated fatty acids, which meets the definition of an UVCB substance based on the analytical characterization. It is insoluble in water (mean 7.4 µg/L at 20°C, pH 6.3 , Schwarzkopf, 2015) with a molecular weight of 399 and 928 g/mol, a log Pow >7.84 based on QSAR predictions (Müller, 2015) and a vapour pressure of <0.0001 Pa at 20 °C (Erler, 2015).

Absorption

Absorption is a function of the potential for a substance to diffuse across biological membranes. The most useful parameters providing information on this potential are the molecular weight, the octanol/water partition coefficient (log Pow) value and the water solubility. The log Pow value provides information on the relative solubility of the substance in water and lipids (ECHA, 2012).

Oral

The smaller the molecule, the more easily it will be taken up. In general, molecular weights below 500 g/mol are favourable for oral absorption (ECHA, 2012). The molecular weight of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate ranges between 399 and 928 g/mol. A molecular weight of 500 g/mol is considered as limit above which oral absorption is assumed to be low. Thus, based on a molecular weight range below and above this “threshold limit”, absorption of especially the smaller components cannot be excluded whereas absorption of the bigger compounds may be low. Absorption after oral administration is also unexpected when the “Lipinski Rule of Five” (Lipinski et al. (2001), Ghose et al. (1999)) is applied to the substance 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate.

The log Pow > 7.34 and the insolubility in water of the substance 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate suggests that the absorption of the highly lipophilic substance may be limited by the inability to dissolve into gastrointestinal (GI) fluids but may be enhanced by micellar solubilisation, as this mechanism is of importance for highly lipophilic substances (log Pow >4), who are poorly soluble in water (1 mg/L or less). In the gastrointestinal tract (GIT), metabolism prior to absorption via enzymes of the microflora may occur. In fact, fatty acid esters with glycerol (glycerides) are hydrolysed by ubiquitously expressed esterases after oral ingestion and are almost completely absorbed (Mattsson and Volpenhein, 1972a). In general, it is assumed that the hydrolysis rate varies depending on the fatty acid chain length and grade of esterification (Mattson and Volpenhein, 1969; Mattson and Volpenhein, 1972a,b)A lower rate of enzymatic hydrolysis in the GIT was shown for compounds with more than 3 ester groups (Mattson and Volpenhein, 1972a,b). The in vitro hydrolysis rate of a pentaerythritol ester was about 2000 times slower in comparison to glycerol esters (Mattson and Volpenhein, 1972a,b).

Moreover in vivo studies in rats demonstrated the incomplete absorption of the compounds containing more than three ester groups. This decrease became more pronounced as the number of ester groups increased, probably the results of different rates of hydrolysis in the intestinal lumen (Mattson and Volpenhein, 1972c).

The available data on oral toxicity of structurally related substances are also considered for assessment of oral absorption. Acute oral toxicity studies were available for Fatty acids, C16-18 and C18-unsatd., branched and linear ester with trimethylolpropane (CAS 403507-18-6) and Fatty acids, C16-18 (even numbered) and C18-unsatd., branched and linear, di and triesters with trimethylolpropane (Formerly CAS No. 85005-23-8). In both studies no signs of systemic toxicity were seen in rats at a concentration of up to 2000 mg/kg bw (Sanders, 2002; Busschers, 1997).

A 90-day oral feeding toxicity study with Fatty acids, C16-18 and C18-unsatd., branched and linear ester with trimethylolpropane (CAS 403507-18-6) displayed no toxicologically relevant effects and therefore the NOAEL was set as > 1000 mg/kg bw/day (McRae, 2004).

The above described studies show that different analogue substances structurally related to 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate revealed a low potential for toxicity after acute and repeated exposure, although no assumptions can be made regarding the absorption potential based on the experimental data. 

In general, after oral ingestion, aliphatic esters of polyhydroxy alcohols (Polyol) and fatty acids are expected to undergo chemical changes in the gastro-intestinal fluids as a result of enzymatic hydrolysis. Thus, 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is assumed to release trimethylolpropane (TMP, parental polyol) and the fatty acid moieties, even if hydrolysis is not assumed to be rapid for polyol esters with more than 3 ester groups. Moreover, pentaerythriol- and dipentaerythritol-esters are expected to be slowly hydrolysed based on in vitro studies, in which the hydrolysis rate of the polyol ester Pentaerythritol tetraoleate was very slow compared to the hydrolysis rate of the triglyceride Glycerol trioleate (Mattson and Volpenhein, 1972). Thus, since it is assumed that esters of polyols (pentaerythritol, dipentaerythritol and trimethylolpropane) have the same metabolic fate, TMP Polyol esters are expected to be hydrolysed slowly as well.

The physico-chemical characteristics of the cleavage products (e.g. physical form, water solubility, molecular weight, log Pow, vapour pressure, etc.) will be different from those of the parent substance before absorption into the blood takes place, and hence the predictions based upon the physico-chemical characteristics of the parent substance do no longer apply but also the physico-chemical characteristics of the breakdown products of the ester; the alcohol and the corresponding fatty acid moieties. (ECHA, 2012). However, also for both cleavage products, it is anticipated that they will be absorbed in the gastro-intestinal tract.

The highly lipophilic fatty acids will be absorbed by micellar solubilisation (Ramirez et al., 2001). A study by Mattson and Nolen (1972) determined the absorbability of the fatty acid moiety of oleate esters of alcohols containing one to six hydroxyl groups. The fatty acid moieties (C18:1) of esters containing less than four ester groups were almost completely absorbed. As the number of ester groups was increased (erythritol and pentaerythritol tetraoleate and xylitol pentaoleate) the absorbability of the fatty acids decreased but was still present.

The remaining hydrolysis product, the TMP, being a highly water-soluble substance (> 100 g/L, OECD SIDS), will readily dissolve into the gastrointestinal fluids.

In summary, the above discussed physico-chemical properties of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate and relevant data from available literature on fatty acid esters suggest a low absorption potential of the parent substance after oral ingestion, especially for the components with high-molecular weight . However, 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is expected to be slowly hydrolysed to the respective polyol and fatty acid moieties, which are expected to be well absorbed with the gastro-intestinal tract

Dermal

The smaller the molecule, the more easily it may be taken up. In general, a molecular weight below 100 g/mol favours dermal absorption, above 500 g/mol the molecule may be too large (ECHA, 2012). The molecular weight of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate ranges between 399 and 928 g/mol, relatively close and above the defined limit above which dermal absorption is considered low, 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is considered to exhibit a rather low penetration potential through the skin. If the substance is a skin irritant or corrosive, damage to the skin surface may enhance penetration (ECHA, 2012). As 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is not considered as skin irritating in humans, an enhanced penetration due to local skin damage can be excluded.

Based on QSAR analysis, a very low dermal absorption value for 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate of 1.0E-06 mg/cm2/event was calculated (Danish QSAR Database, 2012) and hence, the substance has a very low predicted potential for dermal absorption.

For substances with a log Pow above 4, the rate of dermal penetration is limited by the rate of transfer between the stratum corneum and the epidermis, but uptake into the stratum corneum will be high. For substances with a log Pow above 6, the rate of transfer between the stratum corneum and the epidermis will be slow and will limit absorption across the skin, and the uptake into the stratum corneum will be slow. The substance must be sufficiently soluble in water to partition from the stratum corneum into the epidermis (ECHA, 2012). As the water solubility of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is as low as 7.4 µg/L and the log Pow is estimated to be close to ( >7.84) or higher than 10 depending on the grade of esterification, dermal uptake is likely to be very low.

The available data on dermal toxicity on the structural related substance Fatty acids, C16-18 and C18-unsatd., branched and linear ester with trimethylolpropane (CAS No. 403507-18-6) are also considered for assessment of dermal absorption. Topical application of 2000 mg/kg bw of this structural analogue did not induce signs of systemic toxicity in rats (Sanders, 2004). Thus, a low potential for dermal toxicity after acute exposure was determined for Fatty acids, C16-18 and C18-unsatd., branched and linear ester with trimethylolpropane, although no assumptions can be made regarding the dermal absorption potential based on the experimental data.

Overall, the insolubility in water, the high molecular weight (>500), the high log Pow values, the fact that the substance is not irritating to skin and the calculated low dermal absorption potential, implies that dermal uptake of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate in humans is considered as very low.

Inhalation

2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate has a low vapour pressure of less than 0.0001 Pa at 20 °C thus being of low volatility. Therefore, under normal use and handling conditions, inhalation exposure and thus availability for respiratory absorption of the substance in the form of vapours, gases, or mists is not expected to be significant.

However, the substance may be available for respiratory absorption in the lung after inhalation of aerosols, if the substance is sprayed. In humans, particles with aerodynamic diameters below 100 μm have the potential to be inhaled. Particles with aerodynamic diameters below 50 μm may reach the thoracic region and those below 15 μm the alveolar region of the respiratory tract (ECHA, 2012).

Lipophilic compounds with a log Pow > 4, that are poorly soluble in water (0.05 mg/L or less) like 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate (CAS No. 25111-05-1) can be taken up by micellar solubilisation. Esterases present in the lung lining fluid may also hydrolyse the substance, hence making the resulting alcohol and fatty acid moieties available for inhalative absorption..However, as discussed above, hydrolysis of fatty acid esters with more than 3 ester bounds is considered to be slow (Mattson und Volpenhein, 1968, 1972a).

An acute inhalation toxicity study was performed with Heptanoic acid, ester with 2,2-dimethyl-1,3-propanediol (CAS No. 68855-18-5) according to OECD Guideline 436. Three rats per sex were exposed for 4 hours to 5.22 mg/L (mean achieved concentration) test substance aerosol by nose only inhalation (Griffiths, 2012). As no mortality occurred throughout the study period the inhalation LC50 value in rats was determined to be >5.22 mg/L.

Overall, a systemic bioavailability of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate and the respective hydrolysis products in humans is considered likely after inhalation but is not expected to be higher than following oral exposure.

Accumulation

Highly lipophilic substances in general tend to concentrate in adipose tissue, and depending on the conditions of exposure may accumulate within the human body. Although there is no direct correlation between the lipophilicity of a substance and its biological half-life, it is generally the case that substances with high log Pow values have long biological half-lives. The high log Pow of > 7.84 implies that 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate (CAS No. 25111-05-1) may have the potential to accumulate in adipose tissue (ECHA, 2012).

However, as absorption of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate (CAS No. 25111-05-1) is considered to be low, the potential of bioaccumulation is assumed to below as well.

Nevertheless, as further described in the section metabolism below, esters of trimethylolpropane and fatty acids will undergo slow esterase-catalysed hydrolysis, leading to the cleavage products trimethylolpropane and the fatty acids (Fatty acids, C18-unsatd).

The log Pow of the first cleavage product trimethylolpropane is -0.47 and it is highly soluble in water (>100 g/L) (OECD SIDS, 2013). Consequently, there is no potential for trimethylolpropane to accumulate in adipose tissue. The other cleavage products, the fatty acids, can be stored as triglycerides in adipose tissue depots or be incorporated into cell membranes. At the same time, fatty acids are also required as a source of energy. Thus, stored fatty acids underlie a continuous turnover as they are permanently metabolized and excreted. Bioaccumulation of fatty acids only takes place, if their intake exceeds the caloric requirements of the organism.

Overall, the available information indicates that no significant bioaccumulation in adipose tissue of the parent substance and the cleavage products is anticipated.

Distribution

Distribution within the body through the circulatory system depends on the molecular weight, the lipophilic character and water solubility of a substance. In general, the smaller the molecule, the wider is the distribution. If the molecule is lipophilic, it is likely to distribute into cells and the intracellular concentration may be higher than extracellular concentration particularly in fatty tissues (ECHA, 2012).

As discusses above, absorption of 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is considered to be low based on its physicochemical characteristics. Thus, based on the low absorption potential, the high molecular weight and the insolubility in water, 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is not considered to be widely distributed within the human body.

Nevertheless, esters of trimethylolpropane and fatty acids will undergo chemical changes as a result of slow enzymatic hydrolysis, leading to the cleavage products trimethylolpropane and the respective fatty acids (as described above).

Trimethylolpropane, a small, water-soluble substance (134.2 g/mol, log Pow -0.47, water solubility >100 mg/L), will be distributed in aqueous fluids by diffusion through aqueous channels and pores. No protein binding and only poor distribution within fatty tissues is reported (OECD SIDS, 2013).

The fatty acid moieties are distributed in the organism and can be taken up by different tissues. They can be stored as triglycerides in adipose tissue depots or they can be incorporated into cell membranes (Masoro 1977).

Overall, the available information indicates that the parent substance is not expected to be widely distributed whereas distribution of the products of hydrolysis, trimethylolpropane and fatty acids is considered as possible.

Metabolism

Esters of fatty acids are hydrolysed to the corresponding alcohol and fatty acid by esterases (Fukami and Yokoi, 2012)In general, it is assumed that the hydrolysis rate varies depending on the fatty acid chain length and grade of esterification (Mattson and Volpenhein, 1969; Mattson and Volpenhein, 1972a,b).The hydrolysis of esterified alcohols with more than 3 ester groups is assumed to be slow which is supported by in vivo studies in rats, in which a decrease in absorption was observed with increasing degree of esterification. For example, for the polyol ester Pentaerythritol tetraoleate ester an absorption rate of 64% and 90% was observed when administered at 25% and 10% of total dietary fat, respectively, while an absorption rate of 100% was observed for glycerol trioleate (Mattson and Nolen, 1972). In addition, it has been shown in-vitro that the hydrolysis rate of Pentaerythritol tetraoleate was lower when compared with the hydrolysis rate of the triglyceride Glycerol trioleate (Mattson and Volpenhein, 1972a).

Depending on the route of exposure, esterase-catalysed hydrolysis takes place at different places in the organism: after oral ingestion, esters of alcohols and fatty acids undergo enzymatic hydrolysis already in the gastro-intestinal fluids. In contrast, substances which are absorbed through the pulmonary alveolar membrane or through the skin enter the systemic circulation directly before entering the liver where hydrolysis will basically take place.

Thus, 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate is considered to be hydrolysed to the corresponding alcohol (trimethylolpropane) and fatty acids (mostly C18 unsaturated) by esterases, even though at a low rate.

The first cleavage products, the fatty acids are stepwise degraded by beta-oxidation based on enzymatic removal of C2 units in the matrix of the mitochondria in most vertebrate tissues. The C2 units are cleaved as acetyl-CoA, the entry molecule for the citric acid cycle. For the complete catabolism of unsaturated fatty acids such as oleic acid, an additional isomerization reaction step is required. The omega- and alpha-oxidation, alternative pathways for oxidation, can be found in the liver and the brain, respectively (CIR, 1987).

The remaining cleavage product trimethylolpropane is easily absorbed and can either remain unchanged or may further be metabolized or conjugated (e.g. glucuronides, sulfates, etc.) (OECD SIDS, 2013)

Excretion

Based on the low absorption potential, the main route of excretion for 2-Ethyl-2-(hydroxymethyl)-1,3-propanediyl dioleate (parental substance) is expected to be via the faeces.

The first product of hydrolysis, the fatty acid moieties will be mainly excreted by expired air as CO₂, stored as lipids in adipose tissue or used for further physiological properties e.g. incorporation into cell membranes (Lehninger, 1970; Stryer, 1996). The remaining product of hydrolysis, trimethylolpropane, may be excreted via the urine either unchanged or as conjugates (OECD SIDS, 2013).

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