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Diss Factsheets

Administrative data

Link to relevant study record(s)

Reference
Endpoint:
basic toxicokinetics in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22 Nov 2012 to 11 Feb 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Objective of study:
other: hydrolysis in digestive fluid simulants
Qualifier:
according to guideline
Guideline:
other: EFSA Note for Guidance for Food Contact Materials Annex 1 to Chapter III MEASUREMENT OF HYDROLYSIS OF PLASTICS MONOMERS AND ADDITIVES IN DIGESTIVE FLUID SIMULANTS (30 Jul 2008)
Deviations:
no
GLP compliance:
not specified
Radiolabelling:
no
Species:
other: The source of the digestive fluid was not specified in the study report.
Strain:
not specified
Sex:
not specified
Details on test animals or test system and environmental conditions:
Not applicable.
Route of administration:
other: mixing
Vehicle:
other: acetonitrile
Details on exposure:
For the hydrolysis investigation the ester was dissolved in acetonitrile. This solution was added to the intestinal-fluid simulant tempered to 37°C. The concentration of acetonitrile in the reaction mixture was about 0.1%. Samples were taken after 0, 1, 2 and 4 hours. A naphthalene solution in acetone was added as an internal standard to the samples and the enzyme was precipitated by the addition of ice-cold acetone. After filtration the acetone was evaporated. The aqueous solution was acidified with 0.1 M hydrochloric acid (pH 1.2) and extracted three times with dichloromethane. After addition of an alkane standard (tridecane) and derivatization with N-Methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA) at 60°C for one hour the concentrated dichloromethane solution was analysed by gas chromatography coupled with a mass spectrometer (GC/MS). Quantification of the ester and the hydrolysis product was performed specifically by external calibration curves.
Duration and frequency of treatment / exposure:
0, 1, 2 and 4 hours
Dose / conc.:
28.43 ppm
No. of animals per sex per dose / concentration:
Not applicable.
The analysis was performed in triplicate.
Control animals:
no
Details on dosing and sampling:
A duplicate of three different concentrations of the alcohol was performed. For the recovery investigations the alcohol was dissolved in acetonitrile. This solution was added to the intestinal-fluid simulant tempered to 37°C. After 4 hours a naphthalene solution in acetone was added as an internal standard to the samples and the enzyme was precipitated by the addition of ice-cold acetone. Work-up and quantification was performed as already described. For the ester Bis-(2-(2-butoxyethoxy)-ethyl)-adipate the recovery was performed also from water as of the quick decrease of the ester in the enzyme solution. Therefore the stock solution of the ester in acetonitrile was added to water, work-up and quantification was done as already described.
Type:
other: Hydrolysis of Bis-(2-(2-butoxyethoxy)-ethyl)-adipate with intestinal-fluid simulant
Results:
sampling time 0 h: 28.43 ppm (100%) Ester, 0 ppm Alcohol
Type:
other: Hydrolysis of Bis-(2-(2-butoxyethoxy)-ethyl)-adipate with intestinal-fluid simulant
Results:
sampling time 1 h: 0.77 ppm (2.72%) Ester, 14.74 ppm Alcohol
Type:
other: Hydrolysis of Bis-(2-(2-butoxyethoxy)-ethyl)-adipate with intestinal-fluid simulant
Results:
sampling time 2 h: 0.75 ppm (2.63%) Ester, 16.11 ppm Alcohol
Type:
other: Hydrolysis of Bis-(2-(2-butoxyethoxy)-ethyl)-adipate with intestinal-fluid simulant
Results:
sampling time 4 h: 0.72 ppm (2.55%) Ester, 15.01 ppm Alcohol
Details on absorption:
Not applicable.
Details on distribution in tissues:
Not applicable.
Details on excretion:
Not applicable.
Metabolites identified:
yes
Details on metabolites:
The alcohol being released during the hydrolysis reaction is Diethylene glycol mono-n-butyl ether, CAS 112-34-5, MW=162.23 g/mol. Per 1 equivalent ester 2 equivalents alcohol are released.

Table. 1: Quantitative results of the hydrolysis reaction analysis

Contact time

Ester

Alcohol

[h]

[ppm]

[%]

[ppm]

0

28.43

100.00

0.00

1

0.77

2.72

14.74

2

0.75

2.63

16.11

4

0.72

2.55

15.01

Table. 2: Mass balance of the ester hydrolysis reaction

Contact time

Ester

Alcohol

[h]

[µmol]

[µmol] calc.

[µmol] exp.

0

0.327

0.000

0.000

1

0.009

0.636

0.545

2

0.009

0.637

0.596

4

0.008

0.638

0.555

* One ester reacts to two alcohols

Table. 3: Results of the recovery experiment

Ester (from pancreatic medium)

Ester (from water)*

Alcohol (from pancreatic medium)

[ppm]

calc.

[ppm]

exp.

Recovery

[%]

[ppm]

calc.

[ppm]

exp.

Recovery

[%]

[ppm]

calc.

[ppm]

exp.

Recovery

[%]

28.43

0.80

2.81

12.88

13.64

105.9

8.71

7.23

83.0

-

-

-

27.60

30.26

109.6

19.36

17.24

89.0

-

-

-

46.00

48.02

104.4

33.88

31.28

92.3

* As of the quick decrease of the ester in the enzyme solution recovery was performed also from pure water

Conclusion:

The result of the pancreatic digestion of Bis-(2-(2-butoxyethoxy)-ethyl)-adipate shows a decrease of about 97% within 1 hour (recovery experiments indicate a far quicker hydrolysis). No further decrease was measured. The mass balance of this reaction is shown in the Table 2; it displays good congruence of expected and measured values.

Conclusions:
Interpretation of results: ≥ 97% of Bis-(2-(2-butoxyethoxy)-ethyl)-adipate is hydrolysed in digestive fluid simulants within 1 hour.

Description of key information

Oral absorption

Based on available data, absorption after oral ingestion is predicted to be limited as hydrolysis in the gastrointestinal tract is expected to occur. Resulting hydrolysis products are expected to be absorbed readily.

Dermal absorption

The high water solubility, the high molecular weight, the high log Pow value and the lack of potential for skin irritation / corrosion indicate that dermal uptake in humans is likely to be low.

Inhalative absorption

A systemic bioavailability in humans after inhalation exposure cannot be excluded, e.g. after inhalation of aerosols with aerodynamic diameters below 15 μm. The absorption rate is not expected to be higher than that following oral exposure.

Distribution and accumulation

The available information indicates that the intact parent compound is not assumed to distribute throughout the body due to hydrolysis. In contrast, wide distribution within the body is expected for the hydrolysis products. However, no significant bioaccumulation of both the parent substance and its anticipated hydrolysis products in adipose tissue is expected.

Metabolism

Esters of dicarboxylic acids and ethylene glycol monobutyl ethers are hydrolysed to the corresponding ethylene glycol monobutyl ethers and dicarboxylic acids by ubiquitously expressed esterases. A major metabolic pathway for linear dicarboxylic acids, such as adipic acid, is β-oxidation for energy generation. In contrast, ethylene glycol monobutyl ethers are further hydrolysed in the liver to yield ethylene glycols and butanol. While the alcohol is initially oxidised to the corresponding acid, further metabolic degradation includes β-oxidation. The main metabolic pathways of ethylene glycols involve conjugation in the liver with glucuronic acid, sulfate, or glutathione.

Excretion

A considerable rate of hydrolysis is expected in the gastrointestinal tract. Thus, the parent substance is considered to be excreted only to a very minor extent. Following hydrolysis of the parent molecule, the adipic acid and the butanol originating from metabolic breakdown of ethylene glycol monobutyl ethers are not expected to be excreted to a significant degree via the urine or faeces but excreted via exhaled air as CO2. Ethylene glycols are predominantly excreted via the kidneys in the urine following conjugation with glucuronic acid, sulfate, or glutathione for ultimate excretion.

Key value for chemical safety assessment

Bioaccumulation potential:
no bioaccumulation potential

Additional information

There are no studies available in which the toxicokinetic behaviour of the target substance reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9) has been investigated. Therefore, in accordance with Annex VIII, Column 1, Item 8.8 of Regulation (EC) No. 1907/2006 (REACH) and with the ‘Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance’ (ECHA, 2017), an assessment of the toxicokinetic behavior was conducted based on relevant available information. This comprises a qualitative assessment of the available substance-specific data on physico-chemical and toxicological properties and taking into account further available information on a structurally related source substance from which data was used for read-across to cover data gaps.

Reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9) is a multi-constituent substance containing bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate as main constituent (75% (w/w)) and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate as minor constituent. It also contains 2-(2-(2-butoxyethoxy)ethoxy)ethanol] as an impurity in amounts of 1% (w/w). The target substance is a liquid with a water solubility of 6.23 g/L (Nyco, 2016). The molecular weight of its main constituent is 522.67 g/mol, whereas the minor constituent has a molecular weight of 566.7 g/mol. The log Pow was estimated to be in the range 2.41 – 2.69 (QSAR, KOWWIN v1.68) and the calculated vapour pressure is < 0.0001 Pa at 20 °C (QSAR, SPARC online v4.6).

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 absorption

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, 2017). As the molecular weight of reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate is between 522.67 and 566.72 g/mol, absorption of the constituents in the gastrointestinal tract after oral intake is in general not favoured. However, absorption after oral administration of the main constituent is anticipated when the “Lipinski’s Rule of Five” (Lipinski et al., 2001; Ghose et al., 1999) is applied. Except for the molecular weight, all rules are fulfilled for the main constituent since it does not contain more than 5 hydrogen-bond donors (N-H or O-H bonds), no more than 10 hydrogen-bond acceptors (N or O atoms) and the log Pow is not greater than 5. Moreover, the log Pow of 2.41 – 2.69 suggests that the constituents of the target substance are favourable for absorption by passive diffusion, as this mechanism is of importance for substances with a moderate log Pow between -1 and 4, unless they are not very hydrophilic (> 10000 mg/L) (ECHA, 2017). Taking these theoretical considerations, absorption of the target substance or at least of its main constituent is considered likely after oral exposure.

The potential of a substance to be absorbed from the gastrointestinal tract (GIT) may be influenced by chemical changes taking place in gastrointestinal fluids, for instance due to metabolism by gastrointestinal flora or by enzymes released into the gastrointestinal tract or by hydrolysis. This is especially relevant for substances with a high solubility in water, as is the case for the target substance. These changes will alter the physico-chemical characteristics of the substance and hence predictions based upon the physico-chemical characteristics of the parent substance may in some cases no longer apply (ECHA, 2017). After oral ingestion, reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9) is anticipated to undergo hydrolysis of the ester bonds by gastrointestinal enzymes (Lehninger, 1998; Mattson and Volpenhein, 1972a, b). In vitro data regarding hydrolysis confirming this assumption are available for the source substance bis(2-(2-butoxyethoxy)ethyl)adipate (CAS No. 141-17-3). The results of the investigation show that about 97% are hydrolysed within one hour, whereas recovery experiments even indicate a far quicker hydrolysis (FABES, 2013). During hydrolysis of bis(2-(2-butoxyethoxy)ethyl)adipate, two equivalents of the alcohol 2-(2-butoxyethoxy)ethanol and one equivalent of adipic acid are released. Due to the strong structural similarity between the source substance and the target substance, a similar behaviour with respect to hydrolysis is expected for reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate as the main difference is the number of ethylene glycol moieties (i.e. 2 in the source substance and 3 or 4 in the main constituents of the target substance). Since the ethylene glycol side chains in both the source and the target substances are highly flexible with freely rotatable ether functions, no significant steric hindrance is introduced by adding 1 or 2 ethylene glycol moieties compared to the source substance bis(2-(2-butoxyethoxy)ethyl)adipate. Therefore, a similar rate of hydrolysis can be expected for the target substance. Whereas adipic acid has a molecular weight of 146.14 g/mol, the molecular weights of 2-(2-(2-butoxyethoxy)ethoxy)ethanol and 3,6,9,12-tetraoxahexadecan-1-ol resulting from the target substance are 206.26 g/mol and 250.34 g/mol, respectively. Substances with a molecular weight below 200 g/mol may even pass through aqueous pores (ECHA, 2017) and hence absorption of the ethylene glycol monobutyl ethers cannot be excluded and is even very likely for adipic acid.

The available data on oral toxicity are also considered for assessment of oral absorption. No treatment-related systemic effects were observed in a combined repeated dose toxicity study with the reproduction/developmental toxicity screening test performed with the target substance (Intox, 2017). Oral administration of male rates for 4 weeks and of female rats for two weeks prior to mating, the variable time to conception, during pregnancy and 13 days after delivery resulted in a No-Observed-Adverse-Effect-Level (NOAEL) ≥ 1000 mg/kg bw/day, corresponding to the highest dose tested. However, it must be noted that the lack of systemic toxicity observed in the repeated dose study can also be attributed to a low degree of absorption. The lack of systemic toxicity is therefore only an indicator rather than a proof of no or a low toxicity after oral exposure.

Overall, a systemic bioavailability of the target substance reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate in humans cannot be excluded. Although its molecular weight does not favour absorption, other physico-chemical parameters and theoretical considerations indicate that absorption can take place. Moreover, the compounds formed after hydrolysis of the parent molecule, i.e. adipic acid, and the ethylene glycol monobutyl ethers 2-(2-(2-butoxyethoxy)ethoxy)ethanol and 3,6,9,12-tetraoxahexadecan-1-ol, are also considered to be absorbed as they are small (molecular weights < 500 g/mol) and fulfil all theoretical considerations of the “Lipinski’s Rule of Five”. Therefore, a systemic bioavailability is likely after oral uptake of the parent substance.

Dermal absorption

Similar to oral absorption, dermal absorption is favoured for small molecules. In general, a molecular weight below 100 g/mol favours dermal absorption, while a molecular weight above 500 g/mol may be considered too large (ECHA, 2017). As the molecular weight of the target substance ranges between 522.67 and 566.72 g/mol, a dermal absorption is considered not likely. Moreover, 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 itself is also slow. The substance must be sufficiently soluble in water to partition from the stratum corneum into the epidermis (ECHA, 2017). As the water solubility of the target substance is 6.23 g/L and the log Pow is estimated to be 2.41 – 2.69, dermal uptake is again considered to be low based on these theoretical considerations.

The low potential for dermal absorption is also supported by QSAR calculations of the dermal absorption rate. Calculations with the Episuite 4.1, DERMWIN 2.02 tool yielded dermal permeability constants Kp of 7.31E-05 cm/h and 4.14E-05 cm/h for the low molecular weight and high molecular weight constituents, respectively. Both calculated permeability constants are considered to reflect a medium low dermal absorption. Based on these values, the substance has a low potential for dermal absorption.

If the substance is a skin irritant or corrosive, damage to the skin surface may enhance penetration (ECHA, 2017). To this regard primary skin irritation studies conducted with the structurally related source substance bis(2-(2-butoxyethoxy)ethyl)adipate (CAS No. 141-17-3) showed only minor erythema (Consumer Products Testing, 1997b). Therefore, also the target substance is not considered to be irritating or corrosive to skin in humans and an enhanced penetration of the substance due to local skin damage can be excluded.

Overall, the calculated low dermal absorption potential, the high molecular weight (> 100 g/mol) and the fact that the substance is not irritating to skin all lead to the conclusion that dermal uptake of reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9) in humans is possible but considered to be limited.

Inhalation absorption

Reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9) 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. Particles deposited in the nasopharyngeal/thoracic region will mainly be cleared from the airways by the mucocilliary mechanism and swallowed (ECHA, 2017).

Compounds of moderate lipophilicity with a log Pow between -1 and 4 and which are moderately soluble in water, can be absorbed directly across the respiratory tract epithelium by passive diffusion (ECHA, 2017). The unhydrolysed parent substance may therefore be absorbed via this mechanism. Although its water solubility is not favourable as it might be too high, a possible absorption cannot be fully excluded. Moreover, esterases present in the lung lining fluid may also hydrolyse the substance, hence making the resulting ethylene glycol monobutyl ethers and adipic acid available for respiratory absorption. The respective metabolites (adipic acid, 2-(2-(2-butoxyethoxy)ethoxy)ethanol and 3,6,9,12-tetraoxahexadecan-1-ol) are then subsequently readily absorbed. Overall, a systemic bioavailability of the target substance and especially of its hydrolysis products in humans is considered likely after inhalation of aerosols with aerodynamic diameters below 15 μm.

Distribution and accumulation

Distribution of a compound within the body through the circulatory system depends on the physico-chemical 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 its extracellular concentration, particularly in fatty tissues. Furthermore, the concentration of a substance in blood or plasma and subsequently its distribution depends on the rates of absorption. Although there is no direct correlation between the lipophilicity of a substance and its biological half-life, it is generally accepted that substances with high log Pow values have long biological half-lives (ECHA, 2017). Applying these theoretical considerations to the target substance reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9) with its log Pow between 2.41 and 2.69, leads to the conclusion that accumulation in adipose tissue is not likely but cannot be excluded in case of continuous exposure.

However, as shown in the available in vitro test with the structurally similar analogue source substance bis(2-(2-butoxyethoxy)ethyl)adipate (CAS No. 141-17-3) and further discussed in the metabolism section below, the target substance is expected to undergo esterase-catalysed hydrolysis, resulting in adipic acid and the ethylene glycol monobutyl ethers 2-(2-(2-butoxyethoxy)ethoxy)ethanol and 3,6,9,12-tetraoxahexadecan-1-ol. Thus, the distribution and accumulation potential of the hydrolysis products is also regarded to be relevant. The log Pow of the first hydrolysis product adipic acid is 0.08 and its water solubility is larger than 10 g/L (Danish QSAR database, 2017). The ethylene glycol monobutyl ethers have log Pow values of 0.02 and -0.26 for 2-(2-(2-butoxyethoxy)ethoxy)ethanol and 3,6,9,12-tetraoxahexadecan-1-ol, respectively, and their water solubility is > 10 g/L (Danish QSAR database, 2017). Consequently, there is no potential for these compounds to accumulate in adipose tissue when the theoretical considerations mentioned above are applied to these compounds.

Overall, the available information indicates that distribution of the hydrolysis products in various body tissues occurs but no significant bioaccumulation of the parent substance in adipose tissue is anticipated.

Metabolism

A similar metabolic fate of both target and source substances can be assumed based on the similar structural features as both substances contain adipic acid which is fully esterified with ethylene glycol monobutyl ethers. While the ethylene glycol monobutyl ether in the source substance contains a single diethylene glycol substructure, the target substance contains triethylene glycol and/or tetraethylene glycol moieties. Thus, the ethylene glycol monobutyl ether substituents in the target substance contain 1 or 2 additional ethylene glycol units. Although the constituents of the target substance are hence larger in size and have a higher molecular weight, it can be assumed that no significant steric hindrance is introduced when compared to the smaller side chains of the source substance. All parts of the ethylene glycol monobutyl ether side chains are freely rotatable due to the fact that neither a ring system nor π-bonds exert any constraints on rotatability.

There is an in vitro study available investigating the hydrolysis of the source substance bis(2-(2-butoxyethoxy)ethyl)adipate in intestinal fluid simulants as described in the EFSA guidelines ‘Note for Guidance for Food Contact Materials’ (FABES, 2013). The data indicate that after oral ingestion, bis(2-(2-butoxyethoxy)ethyl)adipate undergoes hydrolysis of the ester bonds by gastrointestinal enzymes. The results show that about 97% of bis(2-(2-butoxyethoxy)ethyl)adipate are hydrolysed within one hour, whereas recovery experiments even indicate a far quicker hydrolysis. The experimental result is consistent with the data published by Fukami and Yokoi (2012) which demonstrates that esters of fatty acids are hydrolysed to the corresponding alcohol and fatty acid by esterases. 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 stepwise enzymatic hydrolysis already in the gastrointestinal fluids. In contrast, substances 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. In the first step of hydrolysis of both the target and the source substance, the monoester is produced that is further hydrolysed to the respective ethylene glycol monobutyl ethers and adipic acid.

The metabolic fate of the first hydrolysis products originating from the target substance, 2-(2-(2-butoxyethoxy)ethoxy)ethanol (i.e. triethylene glycol monobutyl ether) and 3,6,9,12-tetraoxahexadecan-1-ol (i.e. tetraethylene glycol monobutyl ether), is anticipated to be similar to that of the well examined compound 2-(2-butoxyethoxy)ethanol (i.e. diethylene glycol monobutyl ether) which is the hydrolysis product resulting from the source substance. Metabolism of ethylene glycol ethers takes place predominantly in the liver where mixed-function oxidases cleave the ether linkage, yielding an ethylene glycol chain and the respective starter alcohol, i.e. butanol in the case of the source and the target substances. Butanol may be consumed in intermediary metabolism to CO2 and water (OECD SIDS, 2006). Ethylene glycols, on the other hand, are supposed to undergo conjugation reactions. This assumption is supported by data obtained from a subchronic repeated dose toxicity study with rats. The results of the study indicate increased liver metabolism after administration of 2-(2-butoxyethoxy)ethanol in drinking water (Johnson et al., 2005). At study termination, rats given 1000 mg/kg bw/day showed increased liver metabolic enzyme activation, including the hepatic Phase II enzyme UDP-glucuronosyltransferase (UGT) together with increased relative liver weights. This finding indicates that glucuronidation is the ultimate metabolic fate of ethylene glycol ethers.

The second hydrolysis product, adipic acid, is metabolized by β-oxidation to succinic and acetic acid and further metabolites (HSDB, 2017). β-oxidation is the degradation pathway of fatty acids based on enzymatic removal of C2 units in the matrix of the mitochondria in most vertebrate tissues. The C2 units are removed from the fatty acid as acetyl-CoA, the entry molecule for the citric acid cycle. The ω- and α-oxidation, alternative pathways for oxidation, can be found in the liver and the brain, respectively (CIR, 1987). Further details of adipic acid metabolism in the rat was reported by Rusoff et al. (1960). Radioactively labelled adipic acid was fed to fasted rats, and the metabolites detected in the urine were analysed as urea, glutamic acid, lactic acid, β-ketoadipic acid, and citric acid, as well as adipic acid. Results from this study also indicate that adipic acid is metabolised by β-oxidation.

The potential metabolites following enzymatic degradation of the target substance were also predicted using the QSAR OECD Toolbox, version 4.1 (OECD, 2017). This QSAR tool predicts the primary and secondary metabolites of the parent compound that may result from enzymatic activity in the liver, in the skin and by the micro-flora in the GIT. Between 34 and 57 hepatic metabolites and 2 – 6 dermal metabolites, depending on the exact structure taken for the prediction, were calculated for the two constituents of reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate (EC No. 943-330-9). As can be expected, the typical hepatic metabolic transformation of the parent compound is the cleavage of the ester bonds yielding either the mono-esters or adipic acid and the ethylene glycol monobutyl ethers in case both ester functions are split. Moreover, fragments originating from cleavage of the various ethylene glycol ether bonds as well as partly oxidised structures, which are hydroxylated at different positions in their ethylene glycol ether chains, are predicted. Metabolites formed in the skin are hydrolysis products only, i.e. mono-esters, adipic acid and the different ethylene glycol monobutyl ethers. No oxidative processes (addition of a hydroxyl group at any position) are predicted to occur in skin. Between 41and 64 metabolites were predicted to result from all kinds of microbiological metabolism for the two constituents considered. This rather high number includes many minor variations in the C-chain length and number of carbonyl and hydroxyl groups, reflecting the diversity of many microbial enzymes identified. Not all of these reactions are expected to take place in the human GIT. In conclusion, the results of the OECD Toolbox simulation support the information on metabolism routes retrieved in the literature.

Overall, the constituents of reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate are expected to be hydrolysed and the hydrolysis products are further metabolised by β-oxidation or conjugation with glucuronic acid, sulfate or glutathione.

Excretion

As a consequence of the enzymatic hydrolysis anticipated for reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate, it is considered to be excreted only to a minor extent. After hydrolysis, the main route of excretion of the various metabolites is expected to be via expired air as CO2 after metabolic degradation (β-oxidation) and via the kidneys in the urine. The metabolism products of reaction mass of bis[2-[2-(2-butoxyethoxy)ethoxy]ethyl]adipate and [2-[2-(2-butoxyethoxy)ethoxy]ethyl](3,6,9,12-tetraoxahexadecyl)adipate that are produced predominantly in the liver may be consumed in intermediary metabolism to CO2 and water, with CO2 ultimately being excreted in expired air. Alternatively, the alcohol and the intermediate ethylene glycol ethers may be conjugated in the liver with glucuronic acid, sulfate, or glutathione for ultimate excretion, predominantly in the urine (OECD SIDS, 2006). Beside excretion after metabolic degradation, adipic acid also can be found unchanged in the urine (Rusoff et al., 1960, HSDB, 2017). Thus, renal excretion after glucuronidation and exhalation as CO2 are the most relevant routes of excretion of the parent substance and its metabolites.

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