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Description of key information

The oral chronic NOAEL was 200 mg/kg bw  in male and female mice in a 2-year carcinogenicity study, based on organ weight changes and mortality at higher doses. The oral subchronic NOAEL was 250 mg/kg bw in male and female Fischer F344 rats and B6C3F1 mice; based on local irritation of the forestomach and increased relative organ weights (stomach, liver in rats and mice;  additionally brain, kidneys, testes in rats).
No treatment-related effects were noted in a OECD Guideline 413 study (Subchronic Inhalation Toxicity: 90-Day). The NOAEC was 120 ppm, i.e. 638.4 mg/m³ the highest concentration tested.
No reliable study with repeated dermal application is available.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
200 mg/kg bw/day
Study duration:
chronic
Species:
mouse
Quality of whole database:
Reliable chronic and subchronic studies with rats and mice are available. These studies are GLP conform and are of high reliability (Klimisch score 1).

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Dr. K. Thomae GmbH, Biberach, Germany
- Age at delivery: 7 weeks
- Weight at study initiation: males 238 (+/- 2.3) g; females 238 (170 +/- 2.2) g
- Housing: singly in wire cages
- Diet: e.g. ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days acclimation to the inhalation chamber


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12 hrs light/dark


IN-LIFE DATES: From: day1 To: day 93
Route of administration:
inhalation
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Remarks on MMAD:
MMAD / GSD: vapour exposure, therefore MMDA not relevant
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: horizontal-flow whole-body inhalation chamber (glas/steel construction, volumes approx. 1.1 m³)
- Method of holding animals in test chamber: rats were kept individually in wire cages
- Test atmosphere generation: For test groups the test substance was delivered to heated evaporators (maintained at 46.4-50.4 °C) by mean of metering pumps. The 40 and 120 ppm test atmosphere was generated using an additional two-component atomizer. In all cases the warmed air was mixed with compressed air (overall stream) and delivered to the inhalation chamber.
- Source and rate of air: compressed air; variable rate
- Temperature, humidity, pressure in air chamber: 23.1 to 23.8°C; positive pressure 10.1 Pascal (control groups), negative pressure -10.2 Pascal (treated groups); 41.8 to 46.2% relative humidity
- Air flow rate: not reported in publication
- Air change rate: not reported in publication
- Treatment of exhaust air: not reported in publication

TEST ATMOSPHERE
- Brief description of analytical method used: samples were analyzed at intervals of about 15 min by gas chromatography (GC-FID; column 1mx22 mm with 10% Triton x 305 on Supelcoport, 102/120 mesh; oven temperature 120 °C; c15-paraffin as internal standard)
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples were analyzed at intervals of about 15 min by gas chromatography (GC-FID; column 1mx22 mm with 10% Triton x 305 on Supelcoport, 102/120 mesh; oven temperature 120 °C; c15-paraffin as internal standard)
Duration of treatment / exposure:
90 days
Frequency of treatment:
6 hours/day, 5 days/week (total of 65 exposures)
Remarks:
Doses / Concentrations:
0, 15, 40 and 120 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
15 (SD 0.67), 39.9 (SD 1.33), 120 (SD 4.8) ppm
Basis:
analytical conc.
No. of animals per sex per dose:
10
Control animals:
yes
Details on study design:
Post-exposure period: none
Positive control:
not required
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS:
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: yes
- Time schedule: pre-treatment and at termination

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION:
- Food consumption: no data contained in the publication; not required

FOOD EFFICIENCY:
- Body weight gain: yes, but no data contained in the publication; not required

WATER CONSUMPTION: no data contained in the publication; not required

OPHTHALMOSCOPIC EXAMINATION: yes
- Time schedule for examinations: pre-treatment and at termination
- Dose groups that were examined: all animals

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at termination, day 94 of the study
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all animals
- Parameters: white and red blood cells, hemoglobin, hematocrit, mean corpuscular volume. mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at termination, day 94 of the study
- Animals fasted: No data
- How many animals: No data
- Parameters: sodium, potassium, chloride, glucose, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, clotting time

URINALYSIS: No data

OTHER:
- cyanide-insensitive palmitoyl-CoA oxidation in liver homogenates
- gross pathology of all animals at termination; determination of organ weights (lungs, liver, kidneys, adrenal glands, and testes)
- histopathology of organs/tissues required by guidelines, and all gross lesions
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Statistics:
- Mean values +/- standard deviation: body weight, body weight gain, hematological and clinical biochemistry parameters, absolute and relative organ weights.
Dunnett's test: comparison of exposure groups with the control group.
Analysis of variance subsequent to Dunnett's test: body weight, body weight gain, hematological and clinical biochemistry parameters
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
1) Analysis of the daily inhalation chamber concentrations revealed that the values obtained closely fitted with the desired nominal level

2) There were no effects regarding the issues below noted at any dose level:

CLINICAL SIGNS AND MORTALITY

BODY WEIGHT AND WEIGHT GAIN

OPHTHALMOSCOPIC EXAMINATION

HAEMATOLOGY

CLINICAL CHEMISTRY

ORGAN WEIGHTS

GROSS PATHOLOGY

HISTOPATHOLOGY: NON-NEOPLASTIC

HISTOPATHOLOGY: NEOPLASTIC (if applicable)


Dose descriptor:
NOAEC
Effect level:
120 ppm (analytical)
Sex:
male/female
Basis for effect level:
other: overall effects
Dose descriptor:
NOAEC
Effect level:
638.4 mg/m³ air (analytical)
Sex:
male/female
Basis for effect level:
other: overall effects
Critical effects observed:
not specified

Under the conditions of the test no treatment-related toxic effects were found in male and female Wistar rats which were exposed to 2-ethylhexanol vapour up to 120 ppm. According to the study authors the concentration of 120 ppm (638.4 mg/m3) corresponds to the calculated saturated vapour concentration at 20°C. However using the highly reliable and newly available vapour pressure of 0.93 hPa a saturation vapour concentration of up to 4884 mg/m³ can be calculated. Concerning the fact that this study did not include test concentrations up to saturation concentration and no toxic effect was observed, the systemic inhalation hazard arising from exposure to 2 -EH might be overestimated when using the NOAEC of 120 ppm.

Conclusions:
No adverse effects were seen in this 90 day inhalation toxicity study (OECD TG 413) in male and female WIstar rats tested up to concentrations of 120 ppm (which was equivalent to saturation at 20°C according to the study authors). Thus the NOAEC of this study was 120 ppm, i.e. 638.4 mg/cubicmetre.
Executive summary:

No treatment-related effects were noted in a OECD Guideline 413 study (Subchronic Inhalation Toxicity: 90-Day) conducted under GLP conditions and using male and female Wistar rats (10 rats per sex and dose). Exposure levels were 0, 15, 40, and 120 ppm. As there were no effects compared with the control groups in either sex on body weight or body weight gain, clinical signs of toxicity and mortality, hematological and clinical biochemistry parameters, ophthalmological parameters, absolute or relative organ weights including testes, or cyanide-insensitive palmitoyl-CoA oxidation as a parameter for hepatic peroxisome proliferation, the NOAEC was 120 ppm, ie. 638.4 mg/m³ (Klimisch, 1998).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
638.4 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
The study is GLP conform and is of high reliability (Klimisch score 1).

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
53.2 mg/m³
Study duration:
subacute
Species:
other: human
Quality of whole database:
The study is reliable (Klimisch score 2) and provides relevant data for humans with the most relevant effect level. Other reliable studies show comparable results for sensory irritation after exposure to the submission substance.

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

Oral route:

Various subacute studies (up to 21 days of treatment) in mice and rats were conducted, mostly as range-finding studies, therefore comprising a smaller range of investigated endpoints. These studies revealed signs of systemic toxicity (e.g. mortality, reduced body weight gain, increased liver weights or increased CN-insensitive palmitoyl coenzyme A activity) at higher dose levels (i.e. at least > 330 mg/kg bw/day) or local irritation of the forestomach after gavage administration (observable at dose levels of approximately 100 mg/kg bw/day, which do not cause systemic effects).

Two reliable subchronic oral gavage rodent studies on 2-EH are available, which were conducted similar to OECD TG 408 (i.e. EPA OTS 798.2650 (90-Day Oral Toxicity in Rodents)) and under GLP. They are reported in the publication of Astill (1996, rat and mouse) and in two study reports of BASF (BASF, 1991, rat, 3 month and BASF, 1991, mouse, 3 month). The systemic toxicity was low in B6C3F1 mice and moderate in Fisher F344 rats. Local irritation caused forestomach inflammation in rats (and in mice, albeit the incidence and the severity were lower compared to rats). 2-EH was a peroxisome proliferator in rats, but not in mice. The NOEL was 125 mg/kg bw/day in rats and mice. The NOAEL is estimated to be 250 mg/kg bw/day. Key findings of these studies are outlined below.

(1) Rat study,90-day oral gavage study using male and female Fischer F344 rats (10 animals per sex and dose) (Astill, 1996 and BASF, 1991, rat, 3 month). The test dose levels were 0, 25, 125, 250, and 500 mg/kg bw/day, based on the results of preliminary 11-day studies.Key results include:

  • Mortalities or clinical findings were not different from controls
  • Food consumption was comparable to controls
  • Body weight gain was reduced in the high dose groups, resulting in decreased terminal weights in males (-7%) and females (-6%); (both p<0.01)
  • Clinico-chemical changes were seen in high dose groups (males: total protein and albumin -13%; females: serum cholesterol -16%); biological significance is unclear
  • Hematology: reticulocytes were increased (25%) in high dose males and females
  • Organ weights: target organs were liver, forestomach, and kidneys; based on increased relative weights (p<0.01) in male and female groups at 250 and 500 mg/kg bw/day. No weight changes were noted at 25 and 125 mg/kg bw/day.
  • Reproductive organs: relative weight of testes was increased, and that of ovaries decreased, at 250 and 500 mg/kg bw/day.
  • Histopathology revealed changes only in high dose animals. Predominantly inflammatory changes in the forestomach which are attributable to the irritation properties of 2 -EH
  • 2 -EH at 500 mg/kg bw/day caused peroxisome proliferation, as evidenced by a statistically significant increase of the hepatic cyanide-insensitive palmitoyl coenzyme A activity in male (p<0.001) and female (p<0.05) rats in Week 13.
  • The subchronic NOEL (based on slight changes of relative organ weights) was 125 mg/kg bw/day in male and female rats.The estimated NOAEL is 250 mg/kg bw/day.

(2) Mouse study,a 90-day oral gavage study using male and female B6C3F1 mice (10 animals per sex and dose) (Astill 1996 and BASF, 1991, mouse, 3 month). The test dose levels were 0, 25, 125, 250, and 500 mg/kg bw/day, based on the results of preliminary 11-day studies.Key results include:

  • There were no mortalities or clinical findings differing from controls
  • Food consumption was comparable to controls
  • Body weight gain was comparable to controls
  • There were no clinico-chemical or hematological changes at any dose level
  • Organ weight changes: target organs were liver and forestomach; a dose-related increase of relative weight was seen in males and females; a level of statistical significance (p<0.05) was gained in males at 250 and 500 mg/kg bw/day
  • Male and female reproductive organs: no weight change noted
  • Histopathology revealed a low incidence of inflammatory changes only in high dose animals; regarded to be incidental; possibly attributable to the irritation properties of 2-EH
  • 2-EH was not a peroxisome proliferator in B6C3F1 mice at up to and including 500 mg/kg bw/day
  • The subchronic NOEL (based on slight changes of relative organ weights) was 125 mg/kg bw/day in male and female rats.The estimated NOAEL is 250 mg/kg bw/day.

Based on these results, the dose for subsequent carcinogenicity studies was set at 50 mg/kg bw/day for the absence of treatment-related effects in rats and mice, and 500 and 750 mg/kg bw/day, respectively, in rats and mice as high doses producing minimal toxicity without altering the life span.

Detailed results of these chronic carcinogenicity studies in rats and mice are presented in section on carcinogenicity (IUCLID section 7.7). From these chronic studies a NOAEL for systemic toxicity of 200 mg/kg bw/d was derived for mice (based on increased mortality, reduced body weights, hepatotoxicity in the high dose group) and 150 mg/ kg bw/d for rats (based on increased mortality in the high dose group).

Dermal route: no valid study identified.

There are two subacute studies of low reliability, due to short exposure periods and insufficient reporting.

In the first study, groups of 10 male and 10 female F-344 rats per dose were dermally exposed to 2-ethylhexanol (undiluted, occluded, at doses of 0, 0.5, or 1.0 ml/kg/day (corresponding to 0, 417, or 834 mg/kg bw/day)). Exposure was for 6 hours per treatment day. Within the experiment the animals were exposed 9 times within 11 days.

There were no treatment-related effects on clinical signs of toxicity, food consumption, or body weight following cutaneous exposure to 2-ethylhexanol. Lymphopenia and decreased spleen weight for high dose females and increased triglycerides for females at both dose levels compared to controls were observed. No other treatment-related effects on clinical pathology measurements or organ weights were observed for males or females at either dose level.Treatment-related anatomic and histologic lesions observed following cutaneous exposure to 2-ethylhexanol were restricted to the site of application and included exfoliation, acanthosis, hyperkeratosis, eschar formation, dermatitis and edema (EPA, 1988).

In the second study only male rats were used (10 males/dose group). Undiluted test substance (2.0 ml/kg/day = 1666 mg/kg bw/day of 2-ethylhexanol, technical grade) was repeatedly (12 treatments) applied to the shaved dorsal skin. Application was non–occlusive; the animals were immobilized for two hours after the application.

On the 10th day a slight reddening and crusting of the skin was evident. The body weights on the 9th and 10th day, and the relative and absolute thymus weights on the 17th day, were significantly reduced. There were no effects on the weights of the heart, liver, spleen, or kidney, the level of protein, albumin, alpha-1-,beta-1-, and gamma-globulin content in serum. 

Histologically the following effects on organs were observed (with at least 3 out of 5 treated animals differing from the controls): 

- Liver: histiocytic and inflammatory granulomas, peripheral fine-droplet fatty degeneration.

- Lungs: interstitial pneumonia, bronchiectasis, severe round-cell bronchitis.

- Kidneys: Epithelial-cell necrosis, cysts, basophilic "ballon nuclei".

- Heart: inter- and intracellular oedema, necrobiotic muscle fibres, interstitial oedema.

- Testes: interstitial oedema, reduced spermiogenesis.

- Thymus: increased "colliodocytes".

- Adrenals: Cortex very rich in lipoids. 

Histochemical investigation of the liver showed increased succinate-dehydrogenase activity and reduced lactate dehydrogenase activity. Tests on acid phosphatase and non-specific alpha-naphtylacetate esterase activity and on fat coloration gave no indications of any changes (Schmidt, 1973).

 

Inhalation exposure:

No treatment-related effects were noted in an OECD Guideline 413 study (Subchronic Inhalation Toxicity: 90-Day) conducted under GLP conditions and using male and female Wistar rats (10 rats per sex and dose). Exposure levels were 0, 15, 40, and 120 ppm. As there were no effects compared with the control groups in either sex on body weight or body weight gain, clinical signs of toxicity and mortality, haematological and clinical biochemistry parameters, ophthalmological parameters, absolute or relative organ weights including testes, or cyanide-insensitive palmitoyl-CoA oxidation as a parameter to measure hepatic peroxisome proliferation, the NOAEC was 120 ppm, ie. 638.4 mg/m³(Klimisch, 1998).

 


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
The study with the longest duration (2 years) was chosen.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
One reliable study available

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
Relevant data on sensory irritation investigated in human volunteers (most relevant effect level see Kiesswetter et al., 2005).

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
No reliable study available; none required as reliable data after oral and inhalative adminsitration are available.

Justification for selection of repeated dose toxicity dermal - local effects endpoint:
No reliable study available; none required as reliable data after oral and inhalative adminsitration are available.

Repeated dose toxicity: via oral route - systemic effects (target organ) digestive: liver; digestive: stomach

Justification for classification or non-classification

Justification for non-classification:

(1) Oral exposure

The NOEL was 250 mg/kg bw/day in the oral subchronic rat and mouse studies, which is far beyond the threshold of the EC Directive 67/548 pertaining to severe health hazards resulting from repeated exposure (R48; threshold: oral, rat: <= 50 mg/kg bw/day).

Similarly, there were no adverse effects on target organs noted at concentrations below the threshold set in the GHS (1st revised edition, 2005) pertaining to specific target organ systemic toxicity following repeated exposure (Chapter 3.9; threshold Category 2, oral, rat: 10 -100 mg/kg bw/day)

References:

EC (1967; 28th ATP 2003) Directive 67/548/EC, section 3.2.4

United Nations (2005), GHS (1st revised edition)

Peroxisome proliferation is a mechanism specific for rodents, and is not relevant for human health.

Reference: Directive 67/548/EC, section 3.2.4

(2) Dermal exposure

Presently no valid studies available

(3) Inhalation exposure

The NOAEC was 120 ppm, i.e. 638.4 mg/m³, in a 90 -day GLP guideline study (OECD TG 413) using male and female Wistar rats. 120 ppm was the highest tested dose level. There was no effect regarding clinical signs of toxicity, body weights or organ weights (including testes in male rats), haematological and clinical biochemistry parameters, ophthalmology, gross pathology and histopathology. Moreover, there was no indication of peroxisome proliferation at termination of the study. Consequently, there was no target organ following inhalation exposure, and no classification is required.