Registration Dossier

Administrative data

Description of key information

Based on read-across from 2-ethylhexanol (CAS 104-76-7), which is a hydrolysis product of the target substance, no hazard was identified.
Oral: chronic NOAEL (rat, m/f) = 934.5-1136.5 mg/kg bw/day (based on read-across from 2-ethylhexanol and after correction for differences in molecular weight)

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented publication comparable to guideline study with acceptable restrictions. No neurology, opthalmoscopy and urine analysis performed.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
yes
Remarks:
no neurology, opthalmoscopy and urine analysis performed
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 36-37 days
- Weight at study initiation: Mean body weight ranges at dosing were (male) 105-114 g and (female) 86-97 g.
- Housing: singly in stainless steel wire cages
- Diet: Kliba rats/mice/hamster maintenance diet, "A" 343 Meal, Klingentalmühle AG, Kaiseraugst, Switzerland, ad libitum
- Water: ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
other: Cremophor EL
Details on oral exposure:
Doses were prepared daily by dispersing 2EH in an aqueous solution of Cremophor EL (5 µg/100 mL) by ultra high speed sonication for 1 min. Homogeneity was maintained by magnetic stirring throughout dosing. Dose volumes were 10 mL/kg, based on weekly body weights. Controls were given 5.0 ml/kg of vehicle.
The dose delivery system, established by prior studies in rats and mice (Astill et al., 1993), was oral gavage of an aqueous emulsion of 2EH stabilized by a nonionic surfactant. This system provided the most stable dose formulation and minimized gastrointestinal tract irritation and inflammation.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentrations and homogeneity were checked by gas chromatographic analysis of samples from each dose level periodically.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
daily on 5 consecutive days per week
Remarks:
Doses / Concentrations:
0, 25, 125, 250, and 500 mg/kg
Basis:

No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
In preliminary subacute studies (11 days) rats were given 0, 100, 330, 1000 and 1500 mg/kg bw/d. Mortality occurred at 1000 and 1500 mg/kg bw/d. The only effects observed at 330 mg/kg were increased relative kidney weights in females, inflammatory edema in the forestomach of one female rat, and a decreased thymus size in males and females. The 11-day dose level free of any treatment-related effects was thus 100 mg/kg.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily, or once daily on non-treatment days

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes, weekly

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: on days 29 and 84
- Animals fasted: Yes
- How many animals:
- Parameters checked: leucocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets and differential leucocytes, and reticulocytes

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: on days 29 and 84
- Animals fasted: Yes
- How many animals:
- Parameters checked: ALAT, ASAT, gamma-GT, AP, Glucose, Urea, total protein, albumin, Creatinine, Cholesterol, Triglycerides, total bilirubin, sodium, potassium, calcium, chloride

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

ORGAN WEIGHTS: Yes, at termination
Sacrifice and pathology:
Moribund animals were euthanized; dead and euthanized animals were immediately necropsied and assessed grossly.
GROSS PATHOLOGY: Yes, adrenals, brains, kidneys, livers, stomachs, testes, and ovaries from all animals were weighed, and with other organs and tissues listed in U.S. EPA Health Effects Guidelines (1987b) fixed in 4% formalin.
HISTOPATHOLOGY: Yes, All tissues from high dose and control animals were stained with hematoxylin—eosin and examined microscopically.
Lungs, livers (including gallbladders in mice), spleens, kidneys, stomachs, sternums, femurs, and femur bone marrows were examined microscopically at intermediate dose levels. Skin, eyes, female mammary glands, thigh musculatures, and extraorbital lacrymatory glands were not examined in the absence of signs of toxicity. Livers were also stained with oil red for lipid content and examined microscopically.
Statistics:
Means and standard deviations were calculated for body weights, food and water consumption, clinical pathology results, and organ weights. Values for test groups were compared with controls in the main study by ANOVA followed by Dunnett's test (Dunnett, 1955, 1964).
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
decrease in males and females at 500 mg/kg bw/d
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
at 500 mg/kg bw/d increase of reticulocytes
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
females at 250 mg/kg bw/d decrease in serum ALAT
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
changes at 250 mg/kg bw/d
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
at 500 mg/kg bw/d in forestomach
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
at 500 mg/kg bw/d in liver
Details on results:
CLINICAL SIGNS AND MORTALITY
One female mouse at 250 mg/kg died during treatment; there were no other mortalities or clinical findings differing from controls in rats at any treatment level.

BODY WEIGHT AND WEIGHT GAIN
There was decreased weight gain in male and female rats at 500 mg/kg, starting at Week 4 in males and Week 11 in females, amounting to weight
losses of 7% in males and 6% in females by Week 13.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
There were no differences from controls at any treatment level in food consumption in rats.

HAEMATOLOGY
There was a 25% increase in reticulocyte numbers in male and female rats at 500 mg/kg.

CLINICAL CHEMISTRY
Differences from controls in treated rats were seen mostly at 84 days (data not shown). Females at 250 and 500 mg/kg/d had 30 and 36% decreases in serum ALT activities, respectively. Females at 500 mg/kg had a 16% decrease in serum cholesterol concentration and males at 500 mg/kg had 13% decreases in total protein and albumin concentrations.

ORGAN WEIGHTS
Significant differences from controls in rats were moderate and limited to the brain, kidneys, liver, stomach, and testes at 250 and 500 mg/kg. Male rat relative brain weights inincreased by 6% at 500 mg/kg, male kidney weights by 8% at 250 and 16% at 500 mg/kg, male liver weights by 8% at 250 and 29% at 500 mg/kg, male stomach weights by 11% at 500 mg/kg, and testis weights by 5.5% at 500 mg/kg. Female rat kidney weights increased by 5% at 250 and 6% at 500 mg/kg, female liver weights by 8% at 250 and 15% at 500 mg/kg, and female stomach weights by 6% at 250 and 16% at 500 mg/kg.

GROSS PATHOLOGY
Gross lesions differing from controls in both species were seen at 500 mg/kg only. In rats 2/10 males and 4/10 females exhibited single
or multiple slightly elevated foci in the forestomach. There were no other gross findings in either species.

HISTOPATHOLOGY: NON-NEOPLASTIC
Dose-related findings in rats (data not shown) were limited to the forestomach and liver at 500 mg/kg. There was a generalized acanthosis
of the forestomach mucosa in 1/10 males with ballooning degeneration of the epithelial wall and acanthosis of the forestomach mucosa in 2/10 males and 5/10 females. There was a moderate decrease in hepatic peripheral lobular fatty infiltration in 4/10 males and 2/10 females and adrenal beta-cell hyperplasia in 3/10 female rats.
Dose descriptor:
NOAEL
Effect level:
125 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects on: clinical signs; mortality; body weight; food consumption; haematology; clinical chemistry; gross pathology; organ weights; histopathology;
Dose descriptor:
LOAEL
Effect level:
250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: decreased serum ALAT and changes in organ weights
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises adequate, reliable (Klimisch score 2) and consistent studies from reference substances with similar structure and intrinsic properties. Read-across is justified based on the structural similarity between the source and target substances, as the source substances either comprises a hydrolysis product (CAS 104-76-7) of the target substance or is a structurally related substance (CAS 61788-89-4) to a hydrolysis product of the target substance. Refer to endpoint discussion for further details. The selected study is thus sufficient to fulfil the standard information requirements set out in Annex VIII-IX, 8.6, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for grouping of substances and read-across

There are no data available on the repeated dose toxicity of Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters (CAS 173832-46-7). In order to fulfil the standard information requirements set out in Annex IX, 8.6, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006, read-across from structurally related substances was conducted.

In accordance with Article 13 (1) of Regulation (EC) No 1907/2006, "information on intrinsic properties of substances may be generated by means other than tests, provided that the conditions set out in Annex XI are met.” In particular for human toxicity, information shall be generated whenever possible by means other than vertebrate animal tests, which includes the use of information from structurally related substances (grouping or read-across).

Having regard to the general rules for grouping of substances and read-across approach laid down in Annex XI, Item 1.5, of Regulation (EC) No 1907/2006 whereby substances may be predicted as similar provided that their physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity.

Overview of repeated dose toxicity:

CAS

173832-46-7 (a)

61788-89-4 (b)

104-76-7

Chemical Name

Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters

Fatty acids, C18-unsatd., dimers

2-Ethylhexanol

Molecular weight

973.62
1085.84
1184.02

556.87;

564.93

130.23

Repeated dose toxicity oral

RA: CAS 61788-89-4

RA: CAS 104-76-7

NOAEL female = 855 mg/kg bw/day (female)

NOAEL = 741 mg/kg bw/day (male)

NOAEL = 125 mg/kg bw/day (male/female)

(a) Category members subjected to the REACh  Phase-in registration deadline of 31 May 2013 are indicated in bold font.

Lack of data for a given endpoint is indicated by “--“.

(b) Surrogate substances are indicated in normal font and are precursors/breakdown products of the target substance (i.e. alcohol and fatty acid moieties). Available data on these substances are used for assessment of toxicological properties by read-across on the same basis of structural similarity and/or mechanistic reasoning as described for the present analogue approach.

The above mentioned substances are considered to be similar on the basis of the structural similar properties and/or activities. The available endpoint information is used to predict the same endpoints for Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters (CAS 173832-46-7).

A detailed analogue approach justification is provided in the technical dossier (see IUCLID Section 13).

 

Discussion

Subchronic oral toxicity

CAS 61788-89-4

A GLP-compliant subchronic oral toxicity study with Fatty acids, C18-unsaturated, dimers was performed similar to OECD guideline 408 in male and female Sprague-Dawley rats (Spurgeon and Hepburn, 1993). Twenty animals per sex and group received the test substance ad libitum at dietary levels of 0, 0.1, 1, 5% (w/w for a period of 13 weeks Based on the reported average body weight and food intake data during Week 1-13, the approximate doses were determined to be 0, 74.1, 740.9, 3591.2 mg/kg bw/day for males and ca. 0, 90.5, 854.9, 4085.5 mg/kg bw/day for females, respectively.

No mortalities and no treatment-related clinical signs were observed in any treatment group during the entire study period. The lower food intake during the first four weeks of the study in rats fed the test material at 5% in the diet may reflect an initial reluctance of the rats to eat the diet. However, no treatment-related changes in body weight were observed in treated animals of any dose group compared to control during the entire study period.

Ophthalmoscopic examination at study termination reveal a persistent pupillary membrane in one male animal of the 0.1% treatment group and ocular opacity in one female animal of the 5.0% treatment group. However, these findings were not considered to be treatment-related.

At necropsy, mesenteric lymph nodes were slightly or moderately enlarged in individual rats of all dose levels compared to controls. Caecal contents appeared yellow-green to yellow in rats given 1.0 and 5.0% in the diet, which was attributed to the yellow colour of the test substance. Furthermore, the incidence of uterine fluid distension was slightly increased in rats fed 5.0% of the test substance in the diet. All other macroscopic findings were considered to be incidental and within the range expected for this age and strain of rat.

Statistically significant decreases in absolute and/or relative organ weight of spleen, kidney and liver were mainly observed in males treated with 1.0% and 5.0% in the diet. In females, a statistically significant decrease in relative organ weights of the liver was observed at all dose levels, but absolute liver weight was only decreased in females treated with 0.1% in the diet. However, these changes were not accompanied by any corresponding histopathological findings, and were thus not considered to be of adverse nature.

Plasma alkaline phosphatase activity (ALP) was increased in males and females fed the test material at 1.0% or 5.0%. In addition, alanine aminotransferase (ALT) was increased in male and female rats fed at 5.0%, which correlated with the increase in biliary hyperplasia and sclerosis in animals of the 5.0% treatment group at histopathological examination. A small increase in plasma bilirubin was observed in male rats fed the test material at 5.0% or 1.0%. However, the levels measured were below the sensitivity of the method and were thus considered with caution. A small reduction in plasma calcium was observed in male and female rats fed the test material at 5.0%. Small reductions in both total serum protein and albumin were also observed in male and female rats of the 5.0% treatment group. However, in the absence of any correlating effect, these changes were not considered to be toxicologically relevant.

The plasma lipids cholesterol and triglyceride were reduced in male and female rats in the 5.0% and 1.0% treatment groups, respectively, which may be a result of interference of the test substance with the absorption of lipid and other nutrients from the gut. The reduction in periportal hepatocyte vacuolation seen on histological examination of the liver could correlate with the reduced plasma lipids, indicating some alteration in lipid metabolism, another possible explanation for the plasma lipid, serum protein and calcium changes.

At microscopic examination, a dose-related increase in the incidence of macrophage aggregation in the mesenteric lymph nodes was observed, which correlated with the mesenteric lymph node enlargement noted at necropsy. The incidence and amount of macrophages was also dose-dependently increased in the spleens of both male and female rats. The macrophages of the mesenteric lymph nodes and spleen revealed a golden/dark brown pigmentation in animals of all dose groups. However, there was no evidence of any degenerative effect associated with these histopathological findings.

Increased cortical vacuolation in the adrenals, associated with decreased cytoplasmic rarefaction was probably due to altered steroidogenesis. However, this finding was not accompanied by any evidence of degenerative change in the adrenal. The significance of the reduced extramedullar haemopoiesis in the adrenals was uncertain, but may possibly correlate with the reduction in neutrophil count in females fed the test material at 5.0% and 1.0%. However, all of the changes observed in the adrenal were minor in nature and thus of no toxicological relevance.

In thyroids, an increase in follicular epithelial hypertrophy was observed in female rats fed 5% of the substance in the diet. Since this effect was only slight and not correlated with any other effects, it was not considered to be toxicologically relevant.

A variety of spontaneous changes was also recorded in animals from all dose groups, which were not considered to be a treatment-related, as they commonly occur in laboratory rats of this age and strain.

Based on the results of this study, a NOAEL of 1% (w/w) was derived for Fatty acids, C18-unsaturated, dimers, corresponding to doses of 741 and 855 mg/kg bw/day for male and female Sprague-Dawley rats, respectively.

CAS 104-76-7

The subchronic oral toxicity of 2-ethylhexanol was investigated in Fischer 344 rats in a study performed similar to OECD guideline 408 (Astill, 1996). Groups of 10 male and 10 female rats received the test substance diluted in Cremophor EL once daily on 5 days/week at dose levels of 25, 125, 250, and 500 mg/kg bw/day for a period of 90 days. A similar constituted group of animals was administered the vehicle alone and served as controls. During the study period, no mortalities and no clinical signs of toxicity were observed in rats at any treatment level. At study termination, body weight gain was decreased in male (-6%) and female (-7%) rats compared to controls. No differences in food consumption were noted between treated animals and controls. Clinical chemistry revealed a 30 and 36% decrease in serum ALT activities in females treated with 250 and 500 mg/kg bw/day, respectively. At 500 mg/kg bw/day, a 16% decrease in serum cholesterol concentration in females and 13% decreases in total protein and albumin concentrations in males were observed. Furthermore, the reticulocyte numbers in male and female rats was increased (+25%) at 500 mg/kg bw/day. Moderate, but statistically significant increases in relative organ weights compared to controls were noted in the brain, kidneys, liver, stomach, and testes of male and female rats at 250 and 500 mg/kg bw/day. At necropsy, 2/10 males and 4/10 females exhibited single or multiple slightly elevated foci in the forestomach at 500 mg/kg bw/day. No other gross pathological findings were observed in any of the animals. Histopathological examination revealed dose-related findings in rats, which were limited to the forestomach and liver at 500 mg/kg bw/day. In the forestomach, generalised acanthosis of the mucosa in 1/10 males with ballooning degeneration of the epithelial wall and acanthosis of the forestomach mucosa in 2/10 males and 5/10 females was observed. However, the effects on forestomach were likely to be attributed to gavage administration of the test substance. In the liver, hepatic peripheral lobular fatty infiltration in 4/10 males and 2/10 females and adrenal beta-cell hyperplasia in 3/10 female rats were noted.

Based on the results of this study, the NOAEL for 2-ethylhexanol in male and female Fischer 344 rats was established at 125 mg/kg bw/day.

Inhalation

There are no data available. Due to their physicochemical properties (low vapour pressure), exposure to Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters via inhalation is unlikely.

Dermal

There are no studies available in which potential toxic effects after long-term dermal exposure have been studied. However, the whole body of evidence on the toxicokinetic behaviour and toxicological activity of Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters indicate that both systemic bioavailability and toxic effects are unlikely to occur upon dermal exposure.

Conclusion on repeated dose toxicity

No studies with Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters investigating repeated dose toxicity are available. Oral, dermal or inhalative absorption of the registered substance is deemed unlikely due to its very high MW and physico-chemical properties. As hydrolysis is possible to a limited extend, it will most likely be the cleavage products that are absorbed, thus a worst case approach considering the repeated dose toxicity of the potential hydrolysis products was performed. For the hydrolysis product 2-Ethylhexanol, a NOAEL of 125 mg/kg bw/day was derived in male and female Fischer 344 rats. For Fatty acids, C18-unsaturated, dimers, which exerts a strong structural relationship to the hydrolysis product Fatty acids, C18-unsaturated, trimers, the NOAEL for male and female Sprague-Dawley rats was set at 741 and 855 mg/kg bw/day, respectively.

The lowest long-term NOAEL of 125 mg/kg bw/day for 2-ethylhexanol (molecular weight = 130.23 g/mol) is selected for hazard assessment of Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters (molecular weight = 973.62-1184.02 g/mol). In order to account for differences in molecular weight, full enzymatic hydrolysis of Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters into Fatty acids, C18 unsatd., trimers and 2-ethylhexanol is assumed.

Thus, a resulting estimated long-term NOAEL ranging from 934.5-1136.5 mg/kg bw/day was established for Fatty acids, C18 unsatd., trimers, 2-ethylhexyl esters.

There are no data available on repeated dose toxicity by the inhalation and dermal routes.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Hazard assessment is conducted by means of read-across from structural analogues and surrogates. The selected study is the most adequate and reliable study based on the identified similarities in structure and intrinsic properties between source and target substance and overall assessment of quality, duration and dose descriptor level (refer to the endpoint discussion for further details).

Justification for classification or non-classification

Based on read-across from the surrogate substance, the available data on repeated dose toxicity do not meet the classification criteria according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.