2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate

Endpoint summary

Link to relevant study record(s)

Reference

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well documented study report which meets basic scientific principles.

Principles of method if other than guideline:

An in vitro human skin penetration study was performed, using radiolabelled test substance. The permeation rate and percentage of octyldodecyl stearoyl stearate through human skin over 48 hours was recorded.

GLP compliance:

no

Radiolabelling:

yes

Species:

other: human skin

Strain:

other: not applicable

Type of coverage:

open

Vehicle:

other: safflower oil

Details on study design:

DOSE PREPARATION
- Method for preparation of dose suspensions: 10 g 10% test substance in safflower oil, at a target activity of 100 µCi/g, was prepared. 15.92 mg 14C-octyldodecyl stearoyl stearate was transferred to a 20 mL vial and unlabelled octyldodecyl stearoyl stearate was added to make the total weight to 1.00153 g. Safflower oil (9.00048 g) was added to produce a solution that contained 10.01% octyldodecyl stearoyl stearate with a theoretical activity of 101.3 µCi/g.

APPLICATION OF DOSE:
The test substance dilution was applied to the skin surface at a concentration of 5 mg/cm².

ANALYSIS
- Method type(s) for identification: liquid scintillation counting
- Validation of analytical procedure: the donor activity was measured in 5 samples taken from the bulk application vehicle to ensure content uniformity. The mean measured donor activity was 101.9 µCi/g, and values varied by less than 1.5%.

Details on in vitro test system (if applicable):

SKIN PREPARATION
- Source of skin: four female donors
- Ethical approval if human skin: no, the samples were obtained from cosmetic reduction surgery
- Type of skin: from the abdomen and breast area
- Preparative technique: the subcutaneous fat was removed from the skin samples by dissection using surgical forceps, scissors and scalpel, and the skin was heat-separated (60 °C for 45 seconds) to generate epidermal membranes. Each epidermal membrane was allowed to dry prior to being frozen, and thawed immediately prior to use. The epidermal membranes (supported on filter paper) were placed on the lower house of greased (silicone grease) diffusion cells, the stratum corneum facing the donor chamber. The upper halves of the diffusion cells were added and the assembly fixed together with pinch clamps.
- Membrane integrity check: the integrity of each membrane was assessed prior to the experiment by determining the permeation of ³H2O over 20 minutes. The skin samples showing water permeation rates greater than 2.0 mg/cm²/h (Kp < 0.002 cm/h) were rejected.
- Storage conditions: stored at -20 °C
- Justification of species, anatomical site and preparative technique: the technique provides separation of the dermis from the epidermis at the epidermal basal layer, generating membranes comprising the entire epidermis.

PRINCIPLES OF ASSAY
- Diffusion cell: Franz-type glass diffusion cells within area available for diffusion of about 1.0 cm² were used. The exact diffusion area was known for each diffusion cell.
- Receptor fluid: 6% Volpo N20 in pH 7.4 phosphate-buffered saline (known volume)
- Static system: the contents of the receptor chamber were continuously agitated by small magnetic followers driven by submersible magnetic stirrers
- Test temperature: the receptor chambers were maintained at 37.0 ± 1°C; the skin surface temperature was maintained at 32.0 ± 1°C
- Occlusion: none
- Other: 200 µL samples were taken from each receptor chamber at 4, 8, 12, 24 and 48 h (application time: 0 h). The 14C-content was determined by liquid scintillation counting. The liquid removed from each cell was replaced with temperature-equilibrated fresh receptor medium. At the end of the 48-h test period, radioactivity remaining on the skin surface and in the donor chamber (including the silicon grease) was removed by gently wiping with cotton buds, which were extracted with isopropyl myristate and tetrahydrofuran respectively, and the samples analysed for 14C-content. The skin samples were removed from this diffusion cell and tape-stripped 12 times using D-squame tape. Following the surface wipe and tape-stripping, the remaining epidermal membrane was solubilised using OptiSolve and assayed for 14C-content.

Absorption in different matrices:

- Receptor fluid, receptor chamber, donor chamber (in vitro test system): less than 0.1% of the applied dose (see Table 1 under 'Any other information on results incl. tables')
- Skin preparation (in vitro test system): 4-5% (total amount in the skin, of which 3.021% in epidermis excluding tape strips)
- Stratum corneum (in vitro test system): 1.485% in tape strips

Total recovery:

- Total recovery: 98.7 ± 1.1% applied dose, of which 94.18% from the skin surface (see Table 2 under 'Any other information on results incl. tables')

Remarks on result:

other:

Table 1: Octyldodecyl stearoyl stearate permeation data for all time points following application

Time (h)	µg/cm²	% applied dose
4	0.013 ± 0.005	0.003 ± 0.001
8	0.012 ± 0.003	0.002 ± 0.001
12	0.017 ± 0.006	0.003 ± 0.001
24	0.044 ± 0.004	0.009 ± 0.001
48	0.023 ± 0.005	0.005 ± 0.001

 

Table 2: 48-h distribution data for octyldodecyl stearoyl stearate

Site	% applied dose	µg/cm²
48-h rinse	94.18 ± 1.39	477.4 ± 16.1
SC tape-strips 1-4	1.051 ± 0.162	5.33 ± 0.82
SC tape-strips 5-12	0.433 ± 0.101	2.18 ± 0.50
Remaining epidermis	3.021 ± 0.406	15.12 ± 1.90
Permeated (48 h)	0.005 ± 0.001	0.023 ± 0.005
Total recovery	98.69 ± 1.12	-

 

Conclusions:

Under the conditions of the study, permeation of octyldodecyl stearoyl stearate was very low. Following 48 h exposure to the 10% solution, between 4 and 5% of the applied dose was recovered from within the skin. The majority of the applied dose (94.18%) was recovered from the skin surface.

Description of key information

2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoateis expected to be absorbed at a low rate via the oral and inhalation route, and have very low absorption potential via the dermal route. The ester will be hydrolysed in the gastrointestinal tract and mucus membranes to the respective fatty acid and fatty alcohol, which facilitates the absorption. The hydrolysis products are expected to be readily absorbed via the oral and inhalation route. The absorbed ester will be hydrolysed mainly in the liver. The fatty acid will most likely be re-esterified to triglycerides after absorption and transported via chylomicrons; while the absorbed alcohol is mainly oxidised to the corresponding fatty acid and then to a triglyceride, as described above. The major metabolic pathway for linear and branched fatty acids is the beta-oxidation pathway for energy generation, while alternatives are the omega-pathway or direct conjugation to more polar products. The excretion will mainly be as CO2 in expired air; with a smaller fraction excreted as conjugated molecules in the urine. No bioaccumulation will take place, as excess triglycerides are stored and used as the energy need rises.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Additional information

Basic toxicokinetics

There were no toxicokinetic studies available for 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate. In accordance with Annex VIII, Column 1, 8.8.1, of Regulation (EC) 1907/2006 and with ‘Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance’ (ECHA, 2017), an assessment of the toxicokinetic behaviour of the substance 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate was conducted to the extent that can be derived from the relevant available information. This comprises a qualitative assessment of the available substance-specific data on physico-chemical and toxicological properties according to the Chapter R.7c Guidance document (ECHA, 2017) and the results of an in vitro human skin penetration study. Furthermore, the available information from source substances was taken into account.

The substance 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate is a UVCB. The main constituents have (a) two ester bonds joining a linear C18 fatty acid (stearic acid), a linear C18 fatty acid with a hydroxyl group (hydroxystearic acid) and a branched C20 fatty alcohol (octyldodecanol) or (b) one ester bond bonds joining either a linear C18 fatty acid (stearic acid) or a linear C18 fatty acid with a hydroxyl group (hydroxystearic acid), and a branched C20 fatty alcohol (octyldodecanol). In addition, a trimer unit (fatty acids and fatty alcohols, three ester bonds), and to a lesser extent oligomer, will be present. The dimer, trimer and oligomers are bulky molecules that will be poorly metabolised due to steric hindrance.

Physico-chemical properties

2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate has the molecular weight 565.01 g/mol (monomer unit) to 1129.93 g/mol (trimer unit). It is a liquid at 20 °C with melting point ≤ - 10 °C, and water solubility of < 0.05 mg/L at 20 °C. The log Pow was estimated to be > 10 and the vapour pressure was calculated to be < 0.0001 Pa at 20 °C.

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, 2017).

Oral

In general, molecular weights below 500 and log Pow values between -1 and 4 are favourable for absorption via the gastrointestinal (GI) tract, provided that the substance is sufficiently water soluble (> 1 mg/L). Lipophilic compounds can be taken up by micellar solubilisation by bile salts, but this mechanism may be of particular importance for highly lipophilic compounds (log Pow > 4), in particular for those that are poorly soluble in water (≤ 1 mg/L) as these would otherwise be poorly absorbed. Solids must be dissolved before absorption; the degree depends on the water solubility (Aungst and Shen, 1986; ECHA, 2017).

Although the physical state favours uptake, some of the physico-chemical characteristics (log Pow, molecular weight and water solubility) of the substance are in a range that indicate poor absorption from the GI-tract following oral ingestion. Therefore, micellar solubilisation is likely be the most relevant absorption mechanism. Due to steric hindrance, the absorption of the dimers and trimers is expected to be negligible.

2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate is predicted to have very low oral absorption due to its physico-chemical characteristics. This prediction is supported by the available acute oral toxicity data. In a study in which rats were administered a single dose of 2 g/kg bw 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate, there was no mortality (key study, 1983). Diarrhoea was observed in all the animals 1 - 4 hours after administration, while 1/5 females had soft stool on Day 2 only. On Day 2, 2/5 females had apparent urinary incontinence, one of these females also showed decreased activity. No effect on body weight and no macroscopic changes were reported. 

In a 28-day repeated dose toxicity study performed with the source substance octyldodecyl isooctadecanoate (CAS 93803-87-3) the NOAEL was considered to be ≥ 1000 mg/kg bw/day, the highest dose level tested (1998, supporting study). Furthermore, no adverse systemic effects were observed in two combined repeated dose toxicity with the reproduction/developmental toxicity screening tests performed with the source substances docosyl docosanoate (CAS 17671-27-1) and tetradecyl oleate (CAS 22393-85-7), up to and including the highest dose level of 1000 mg/kg bw/day (supporting study, 2014; key study, 2014).

The potential of a substance to be absorbed in the GI-tract may be influenced by chemical changes taking place in GI-fluids, for instance as a result of metabolism by GI-flora, by enzymes released into the GI-tract or by hydrolysis. These changes will alter the physico-chemical characteristics of the substance and hence predictions based on the physico-chemical characteristics of the parent substance may in some cases no longer apply (ECHA, 2017). In general, alkyl esters are readily hydrolysed in the GI-tract, blood and liver to the corresponding alcohol and fatty acid by the ubiquitous carboxylesterases. There are indications that the hydrolysis rate in the intestine catalysed by pancreatic lipase is lower for alkyl esters than for triglycerides, which are the natural substrates of this enzyme. The hydrolysis rate of linear esters increases with increasing chain length of either the alcohol or acid. Branching reduces the ester hydrolysis rate, compared with linear esters (Mattson and Volpenhein, 1969, 1972; WHO, 1999). Furthermore, the size of the structure and the two ester bonds may reduce the hydrolysis rate. Based on the generic information on hydrolysis of alkyl esters, the target substance 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate may be enzymatically hydrolysed to hydroxystearic acid, stearic acid and octyldodecanol , assuming both ester bonds are broken. The monomer unit with the single ester bond is predicted to be hydrolysed. The hydrolysis of oligomers is expected to be very limited, due to the steric hindrance.

The fraction of ester that is hydrolysed to acid and alcohol moieties is more likely to be absorbed from the GI-tract than the parent substance. In general, free fatty acids are readily absorbed by the intestinal mucosa. Within the epithelial cells, fatty acids are mainly (re-)esterified with glycerol to triglycerides. Branching is likely to reduce the absorption rate (Greenberger et al., 1966; IOM, 2005; Mattson and Volpenhein, 1964; OECD, 2006; Sieber, 1974). With increasing chain-length the fatty acids are increasingly absorbed via the lymphatic route, and will be metabolised in the liver (Ramirez et al., 2001).

In conclusion, the log Pow, water solubility and molecular weight of 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate suggest that oral absorption is likely to be very low. The monomer (and to some extent the dimer) of the substance is expected to undergo enzymatic hydrolysis in the GI-tract to a certain extent and therefore absorption of the ester hydrolysis products is also relevant. The absorption rate of the hydrolysis products is assumed to be high, as no experimental data is available.

Dermal

The dermal uptake of liquids and substances in solution is higher than that of dry particulates, since dry particulates need to dissolve into the surface moisture of the skin before uptake can begin. Molecular weights below 100 g/mol favour dermal uptake, while for those above 500 g/mol the molecule may be too large. Dermal uptake is anticipated to be low if the water solubility is < 1 mg/L; low to moderate if it is between 1-100 mg/L; and moderate to high if it is between 100-10000 mg/L. Log Pow values in the range of 1 to 4 are favourable for dermal absorption (values between 2 and 3 are optimal), in particular if the water solubility is high. For substances with a log Pow above 4, the rate of penetration may be limited by the rate of transfer between the stratum corneum and the epidermis, but uptake into the stratum corneum will be high. Log Pow values above 6 reduce the uptake into the stratum corneum and decrease the rate of transfer from the stratum corneum to the epidermis, thus limiting dermal absorption (ECHA, 2017).

The target substance 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate has a molecular weight of 565.01 g/mol (monomer unit) to 1129.93 g/mol (trimer unit) and very low water solubility; therefore a low dermal absorption potential might be assumed (ECHA, 2017). The log Pow is > 10, which means that the uptake into the stratum corneum is predicted to be slow and the rate of transfer between the stratum corneum and the epidermis will be slow (ECHA, 2017).

An in vitro human skin penetration study was performed with 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate, using radiolabelled substance (LoA, 2001, supporting study). Under the conditions of the study, permeation of 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate through human skin was very low. Following 48 hours exposure to a 10% solution in safflower oil, 4 - 5% of the applied dose was recovered from within the skin and 0.05% had penetrated the skin. The majority of the applied dose (94.18%) was recovered from the skin surface. This indicates a very low skin penetration will occur in vivo.

If a substance shows skin irritating or corrosive properties, damage to the skin surface may enhance penetration. If the substance has been identified as a skin sensitizer then some uptake must have occurred although it may only have been a small fraction of the applied dose (ECHA, 2017).

The target substance did not cause skin irritation in vivo in the rabbit (key study, 1978). Furthermore, the available data for source substances provide no indications for skin irritating effects in human volunteers (key study, 2001). No skin effects were noted in the acute dermal toxicity study at the limit dose of 2000 mg/kg bw, performed with a source substance (key study, 1998). The result of the skin sensitisation test performed in guinea pigs with a source substance was negative (WoE, 1991). Therefore, no penetration of the substance due to skin damage is expected.

Overall, based on the available information, the dermal absorption potential of 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate is predicted to be very low or negligible.

Inhalation

2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate is a liquid with low vapour pressure (< 0.0001 Pa at 20 °C), and therefore low volatility. Therefore, under normal use and handling conditions, inhalation exposure and availability for respiratory absorption of the substance in the form of vapours, gases, or mists is considered to be limited (ECHA, 2017). However, the substance may be available for inhalatory absorption after inhalation of aerosols, if the substance is sprayed (e.g. as a formulated product). 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. Particles deposited in the nasopharyngeal/thoracic region will mainly be cleared from the airways by the mucocilliary mechanism and swallowed.

Absorption after oral administration of the substance is mainly driven by enzymatic hydrolysis of the ester bond to the respective metabolites and subsequent absorption of the breakdown products. To ensure effective absorption in the respiratory tract, enzymatic hydrolysis in the airways would be required and the presence of esterases and lipases in the mucus lining fluid of the respiratory tract would be important. Due to the physiological function of enzymes in the GI-tract for nutrient absorption, esterase and lipase activity in the lung is expected to be lower in comparison to the gastrointestinal tract. Therefore, hydrolysis comparable to that in the gastrointestinal tract and subsequent absorption in the respiratory tract is considered to happen at a lower rate. The molecular weight, log Pow and water solubility indicate that the substance may be absorbed across the respiratory tract epithelium by micellar solubilisation to a limited extent. However, low water solubility (< 0.05 mg/L) does restrict the diffusion/dissolving into the mucus lining before reaching the epithelium, and it is not clear which percentage of the inhaled aerosol could be absorbed as the ester. 

An acute inhalation toxicity study was performed with the read-across (source) substance 2-ethylhexyl oleate (CAS 26399-02-0), in which rats were exposed nose-only to > 5.7 mg/L of an aerosol for 4 hours (key study, 2010). No mortality occurred and no toxicologically relevant effects were observed which might be due to no/low toxic potential or no/low absorption rate. Therefore, the target substance is not expected to be acutely toxic by the inhalation route.

Due to the limited information available a worst case approach is applied, and absorption via inhalation is assumed to be as high as via the oral route.

Distribution and Accumulation

Distribution of a compound within the body depends on the physicochemical properties of the substance; especially the molecular weight, the lipophilic character and the water solubility. 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, 2017). Following the predicted hydrolysis of the target substance, the resulting hydrolysis products are not lipophilic and are not expected to accumulate in fatty tissue. The fraction of absorbed ester will be hydrolysed in the liver and is therefore also unlikely to accumulate in fatty tissue.

As discussed under oral absorption, 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate will undergo some enzymatic hydrolysis in the gastrointestinal tract prior to absorption. The fraction of ester absorbed unchanged will undergo enzymatic hydrolysis by ubiquitous esterases, primarily in the liver (Fukami and Yokoi, 2012). The distribution and accumulation of the hydrolysis products is considered the most relevant. After being absorbed, linear and simple branched fatty acids are (re-) esterified along with other fatty acids into triglycerides and released in chylomicrons into the lymphatic system. This route of absorption and metabolism of a fatty acid was shown in an in vivo study performed by Sieber (1974). Twenty-four hours after intraduodenal administration of a single dose of [1-¹⁴C]-radiolabelled octadecanoic acid to rats, 52.5 ± 26% of the radiolabelled carbon was recovered in the lymph. A large majority (68 - 80%) of the recovered radioactive label was incorporated in triglycerides, 13 - 24% in phospholipids and 0.7 - 1% in cholesterol esters. No octadecanoic acid was recovered. Almost all the radioactivity recovered in the lymph was localized in the chylomicron fraction. Fatty acids of carbon chain length ≤ 12 may be transported directly to the liver via the portal vein as the free acid bound to albumin, instead of being re-esterified. This is supported by the Sieber study (1974), in which, following the same protocol as described above, administration of hexanoic acid lead to only 3.3% recovery from lymphatic fluid. Chylomicrons are transported in the lymph to the thoracic duct and subsequently to the venous system. On contact with the capillaries, enzymatic hydrolysis of chylomicron triacylglycerol fatty acids by lipoprotein lipase takes place. Most of the resulting fatty acids are taken up by adipose tissue and re-esterified into triglycerides for storage. Triacylglycerol fatty acids are also taken up by muscle and oxidized to derive energy or they are released into the systemic circulation and returned to the liver, where they are metabolised, stored or re-enter the circulation (IOM, 2005; Johnson, 1990; Johnson, 2001; Lehninger, 1993; NTP, 1994; Stryer, 1996; WHO, 2001). Methyl substituted fatty acids were shown to be metabolised via beta- and omega-oxidation and can be expected to have a similar distribution to linear fatty acids (WHO, 1998). Highly branched fatty acids are expected to be widely distributed, although the distribution may be facilitated via other routes than the chylomicrons, due to the steric hindrance. Absorbed alcohols are mainly oxidised to the corresponding fatty acid and then follow the same metabolism as described above for fatty acids, to form triglycerides.

Substances that are absorbed through the pulmonary alveolar membrane or through the skin enter the systemic circulation directly before transport to the liver where hydrolysis will generally take place.

The accumulation of fatty acids takes place only if their intake exceeds the caloric requirements of the organism. The potential of the fatty acids derived from the parent substance to accumulate, is considered to be low.

Metabolism

The metabolism of 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate is expected to initially occur via a stepwise enzymatic hydrolysis of the ester resulting, ultimately, in the corresponding hydroxystearic acid, stearine and octyldodecanol. The esterases catalysing the reaction are present in most tissues and organs, with particularly high concentrations in the GI-tract and the liver (Fukami and Yokoi, 2012).

The fatty acids can be metabolised directly following absorption or following de-esterification of triglycerides. The beta-oxidation pathway for energy generation is the major metabolic pathway for linear and an important pathway for simple branched fatty acids. In this multi-step process, the fatty acids are at first esterified into acyl-CoA derivatives and subsequently transported into cells and mitochondria by specific transport systems. In the next step, the acyl-CoA derivatives are broken down into acetyl-CoA molecules by sequential removal of 2-carbon units from the aliphatic acyl-CoA molecule. Further oxidation via the citric acid cycle leads to the formation of H₂O and CO₂(Lehninger, 1993). Branched-chain acids can be metabolised via the same beta-oxidation pathway as linear, depending on the steric position of the branch, but at lower rates (WHO, 1999). For fatty acids with complex branching, the alpha-oxidation pathway is the major metabolic pathway, as a methyl substituent at the beta-position blocks certain steps in the beta-oxidation (Mukherji, 2003). Generally, a single carbon unit is cleaved off the branched acid in an additional step before the removal of 2-carbon units continues.

A fraction of the octyldodecanol may be subjected to omega-oxidation due to steric hindrance by the methyl groups at uneven position, which results in the formation of various diols, hydroxyl acids, ketoacids or dicarbonic acids. In contrast to the products of beta-oxidation, these metabolites may be conjugated to glucuronides or sulphates.

The potential metabolites following enzymatic metabolism of the test substance were predicted using the QSAR OECD toolbox (OECD, 2016). This QSAR tool predicts which metabolites of the test substance may result from enzymatic activity in the rat, in rat liver with S9-metabolic activation, in the skin, and by intestinal bacteria in the gastrointestinal tract.

Twenty (20) metabolites were predicted as described above in vivo in the rat. For most of these metabolites, both ester bonds remained intact and a hydroxyl group or a methyl group was added to a CH₂- or CH₃- group. In general, the hydroxyl groups make the substances more water-soluble and susceptible to metabolism by phase II-enzymes. In some cases the ester bond was broken, after which the hydrolysis products may be metabolised further. The resulting liver and skin metabolites are the products of alpha-, beta- or omega-oxidation (= addition of hydroxyl group). In the case of omega-oxidation, it is followed by further oxidation to the aldehyde, which is then oxidised to the corresponding carboxylic acid. For a branched fatty acid, the alpha- and omega pathways are particularly relevant.

Twenty (20) hepatic (S9-) metabolites and 4 dermal metabolites were predicted. Primarily, one or both ester bonds were hydrolysed as described above. The metabolites formed in the skin are expected to enter the blood circulation and have the same fate as the hepatic metabolites.  Two hundred and forty-four (244) metabolites were predicted to result from all kinds of microbiological metabolism. The high number includes many minor variations in the c-chain length and number of carbonyl- and hydroxyl groups; reflecting the many microbial enzymes identified. Not all of these reactions are expected to take place in the human GI-tract. The results of the OECD Toolbox simulation support the information on metabolism routes retrieved in the literature.

There is no indication that 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate is activated to reactive intermediates under the relevant test conditions. The experimental studies performed on genotoxicity (Ames test, gene mutation in mammalian cells in vitro, in vivo micronucleus test) using the target and source substances were consistently negative, with and without metabolic activation (Ames, 1998; Ames, 1994; HPRT, 1994; MLA, 2010; MNT, 2010). The result of the skin sensitisation study performed in guinea pigs using a source substance, was likewise negative (key, 1998).

Excretion

In general, linear fatty acids and fatty acids with simple branching derived directly from the hydrolysis of the ester or from the oxidation of the corresponding alcohol, as well as the fatty acids, will be metabolised for energy generation, stored as lipid in adipose tissue (if there is an excess) or used for further physiological functions, like incorporation into cell membranes (Lehninger, 1993). Therefore, the fatty acid metabolites are not expected to be excreted to a significant degree via the urine or faeces but to be excreted via exhaled air as CO₂ or stored as described above. Experimental data with ethyl oleate (CAS 111-62-6, ethyl ester of oleic acid) support this principle. The absorption, distribution, and excretion of 14C-labelled ethyl oleate was studied in Sprague Dawley rats after a single, oral dose of 1.7 or 3.4 g/kg bw. At sacrifice (72 h post-dose), 12- 33% was recovered from the carcass; low concentrations of radioactivity was measured in most of the organs/tissues. The main route of excretion of radioactivity in the groups was via the expired air as CO₂. 12 h after dosing, 40-70% of the administered dose was excreted in expired air (consistent with beta -oxidation of fatty acids). 7-20% of the radioactivity was eliminated via the faeces, and approximately 2% via the urine (Bookstaff et al., 2003).

Glucuronides or sulphates resulting from omega-oxidation will be excreted via urine or bile or cleaved in the gut with the possibility of reabsorption (entero-hepatic circulation) (WHO, 1998).

The fraction of 2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate that is not absorbed in the GI-tract, will be excreted via the faeces. This is also the case for the monomer and oligomer.

References

Bookstaff et al. (2003). The safety of the use of ethyl oleate in food is supported by metabolism data in rats and clinical safety data in humans. Regul Toxicol Pharm 37: 133-148.

ECHA (2017). Guidance on information requirements and chemical safety assessment, Chapter R.7c: Endpoint specific guidance.

Fukami, T. and Yokoi, T. (2012). The Emerging Role of Human Esterases. Drug Metab Pharmacokinet 27(5): 466-477

Greenberger et al. (1966). Absorption of medium and long chain triglycerides: factors influencing their hydrolysis and transport. J Clin Invest. 45(2):217-27.

IOM (2005). Dietary Reference Intakes for Energy, Carbohydrate, Fiber, Fat, Fatty Acids, Cholesterol, Protein, and Amino Acids (Macronutrients). Institute of the National Academies. The National Academies Press. http://www.nap.edu/openbook.php?record_id=10490&page=R1

Johnson, R.C. et al.(1990). Medium-chain-triglyceride lipid emulsion: metabolism and tissue distribution. Am J Clin Nutr 52(3):502-8.

Johnson W. Jr; Cosmetic Ingredient Review Expert Panel. (2001). Final report on the safety assessment of trilaurin, triarachidin, tribehenin, tricaprin, tricaprylin, trierucin, triheptanoin, triheptylundecanoin, triisononanoin, triisopalmitin, triisostearin, trilinolein, trimyristin, trioctanoin, triolein, tripalmitin, tripalmitolein, triricinolein, tristearin, triundecanoin, glyceryl triacetyl hydroxystearate, glyceryl triacetyl ricinoleate, and glyceryl stearate diacetate. Int J Toxicol. 2001;20 Suppl 4:61-94.

Lehninger, A.L., Nelson, D.L. and Cox, M.M. (1993). Principles of Biochemistry. Second Edition. Worth Publishers, Inc., New York, USA. ISBN 0-87901-500-4.

Mattson, F.H. and Volpenhein, R.A. (1964). The digestion and absorption of triglycerides. J Biol Chem. 239:2772-7.

Mattson, F.H. and Volpenhein, R.A. (1969). Relative rates of hydrolysis by rat pancreatic lipase of esters of C2 - C18 fatty acids with C1 – C18 primary n-alcohols.J Lipid Res Vol(10): 271 – 276

Mattson, F.H. and Volpenhein, R.A. (1972). Hydrolysis of fully esterified alcohols containing from one to eight hydroxyl groups by the lipolytic enzymes of the rat pancreatic juice. Journal of Lipid Research 13: 325-328

Mukherji M. et al. (2003). The chemical biology of branched-chain lipid metabolism. Progress in Lipid Research 42: 359-376.

National Toxicology Program (NTP) (1994) Comparative toxicology studies of Corn Oil, Safflower Oil, and Tricaprylin (CAS Nos. 8001-30-7, 8001-23-8, and 538-23-8) in Male F344/N Rats as vehicles for gavage. http://ntp.niehs.nih.gov/ntp/htdocs/LT_rpts/tr426.pdf (2011-12-19). Report No.: C62215. Owner company: U.S. Department of Health and Human Services, Public Health Services, National Institutes of Health.

OECD (2006). Long Chain Alcohols. SIDS Initial Assessment Report For SIAM 22. Paris, France, 18-21 April 2006. TOME 1: SIAR. http://webnet.oecd.org/Hpv/UI/SIDS_Details.aspx?id=7A14361C-4676-4339-A915-2CFD51F12483

OECD (2016). (Q)SAR Toolbox v3.4. Developed by Laboratory of Mathematical Chemistry, Bulgaria for the Organisation for Economic Co-operation and Development (OECD). Prediction performed on 28 March 2017.http://toolbox.oasis-lmc.org/?section=overview

Ramirez, M. et al. (2001). Absorption and distribution of dietary fatty acids from different sources. Early Hum Development 65 Suppl.: S95 – S101

Sieber, S.M., Cohn, V.H., and Wynn, W.T. (1974). The entry of foreign compounds into the thoracic

duct lymph of the rat.Xenobiotica 4(5), 265.

Stryer, L. (1996). Biochemie. 4. Auflage. Heidelberg Berlin Oxford: Spektrum Akademischer Verlag.

WHO (1998). Safety evaluation of certain food additives and contaminants. WHO Food Additives Series 40. Joint FAO/WHO Expert Committee on Food Additives.

WHO (1999). Evaluation of certain food additives and contaminants. Forty-ninth report of the joint FAO/WHO Expert Committee on Food Additives. WHO Technical Report Series 884. ISBN 92 4 120884 8.

WHO (2001). Safety Evaluation of Certain Food Additives and Contaminants: Aliphatic Acyclic Diols, Triols, and Related Substances. WHO Food Additives Series No. 48.