2-ethylhexanoic acid

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, near guideline study, with minor restrictions in design and/or reporting but otherwise adequate for assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OTS 795.2600 (Subchronic Oral Toxicity Test)

GLP compliance:

yes

Remarks:

Health and Environmental Laboratories, Eastman Kodak Company

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-ethylhexanoic acid

EC Number:

205-743-6

EC Name:

2-ethylhexanoic acid

Cas Number:

149-57-5

Molecular formula:

C8H16O2

IUPAC Name:

2-ethylhexanoic acid

Details on test material:

- Purity: 99.9% (±0.05%; GC)
- The test material is stable in open feeders for at least 7 days (GC)

Test animals

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Wilmington, MA
- Age at study initiation: 6 weeks
- Weight at study initiation: males: 23 (± 0.8) g; females: 19 (± 0.8) g
- Housing: individually
- Diet (e.g. ad libitum): Agway Prolab Animal Diet (RMH 3200), certified ground chow; ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 2 weeks


ENVIRONMENTAL CONDITIONS
- Temperature (°F): 68-72
- Humidity (%): 46-67
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
Diets were prepared containing 0.0, 0.1, 0.5 or 1.5% of the test material. The diets were frozen in closed amber glass bottles until used. The test material was stable for at least 35 days in the diets when refrigerated. Stability, homogeneity and concentration analyses were conducted using GC.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyzed concentrations (mean ± SD; GC): 0.091 (± 0.004), 0.45 (± 0.02) and 1.45 (± 0.05)%

Duration of treatment / exposure:

91-93 days

Frequency of treatment:

7 days/week

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
0, 0.1, 0.5 and 1.5%. 28-day recovery groups: 0 and 1,5%
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
0, 0.091, 0.45 and 1.45% (recovery groups: 0 and 1.45%)
Basis:
other: analyzed in diet

Doses / concentrations 3

Remarks:

Doses / Concentrations:
0, 180, 885 and 2728 mg/kg/day for males (recovery: 0 and 2895 mg/kg/day); 0, 205, 1038 and 3139 mg/kg/day for females (recovery: 0 and 3101 mg/kg/day)
Basis:
other: actual dose

No. of animals per sex per dose:

10

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: based on a 14-day feeding study in mice
- Post-exposure recovery period in satellite groups: 28-29 days

Positive control:

Not included

Examinations

Observations and examinations performed and frequency:

CLINICAL OBSERVATIONS: Yes
- Detailed observations: on the mornings of body weight measurement.
- Cage side observations: every workday afternoon and on mornings on which body weights were not collected. Animals were checked for mortality on weekends.

BODY WEIGHT: Yes
- Time schedule for examinations: days 0, 3, 7, and at least once weekly thereafter.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Feed weights (g) were collected on days 3, 7, and at least once weekly thereafter.

OPHTHALMOSCOPIC EXAMINATION: Yes, using an indirect ophthalmoscope after dilation of the pupils with 1% Mydriacyl.
All mice were examinde prior to the start of the study. During the last week of exposure, five male and five female non-recovery animals from each dose level were examined.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at necropsy
- Anaesthetic used for blood collection: Yes (CO2 anaesthesia)
- Animals fasted: overnight
- How many animals: five animals per sex, per dose.
- Parameters: hemoglobin concentration, hematocrit, red blood cell count, white blood cell count, differential white blood cell count, platelet count, red blood cell indices, and examination of the blood smears for cellular morphology.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at necropsy
- Animals fasted: overnight
- How many animals: five animals per sex per dose
- Parameters: Non-recovery groups: aspartate aminotransferase, alanine aminotransferase, total bilirubin, total protein, albumin, creatinine, urea nitrogen, glucose, gamma glutamyl transpeptidase, triglycerides, cholesterol, sodium, potassium, chloride, calcium and phosphorus. Recovery groups: alanine aminotransferase, total bilirubin, urea nitrogen, triglycerides, cholesterol.

URINALYSIS: Yes
In the week prior to termination of exposure, five males and five females from each dose group were place in metabolism cages for 24-hour urine collections. Parameters: specific gravity, osmolality, volume, glucose, bilirubin, ketones, blood, protein, urobilinogen, nitrite, leukocytes and pH.

Sacrifice and pathology:

Organ weights: liver, kidneys, adrenal glands, testes, ovaries, brain.
Histopathology: all non-recovery high-dose and control animals: trachea, lungs, heart, aorta, esophagus, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, pancreas, liver, salivary glands, kidneys, urinary bladder, pituitary gland, adrenal glands, thyroid glands, parathyroid glands, thymus, spleen, mesenteric lymph nodes, bone marrow (femoral), brain (including sections of medulla/pons, cerebellar cortex, and cerebral cortex), sciatic nerve, quadriceps femoris, testes, ovaries, vagina, uterus, fallopian tubes, sternum with bone marrow, and gross lesions.
The liver, kidneys, lungs, target organs, and gross lesions for animals from all dose levels were examined.
Because no signs of toxicity or target organ involvement was observed, no histopathology was performed on cervical/mid-thoracic/lumbar spinal cord, epididymides, male accessory sex glands, male mammary gland, female mammary gland, femur (including articular surface), skin, and exorbital lachrymal glands.
For recovery animals, histopathology was performed on the liver, kidneys, lungs, and gross lesions.

Statistics:

Means were calculated for body weight, feed consumption, and organ weights. Numerical data were evaluated using Bartlett's test, one-way ANOVA, and Duncan's multiple range test.

Results and discussion

Results of examinations

Details on results:

CLINICAL SIGNS AND MORTALITY
No mortality occured during the study.

BODY WEIGHT AND WEIGHT GAIN
A reduced mean body weight was observed in males and females of the high-dose group (5.2% and 13.8% below control weights on day 91, respectively) and in females of the mid-dose group (5.6% below control weights on day 91). At the end of the recovery period, body weight was still 4.7% and 10.1% below control weights in males and females of the high-dose group, respectively.

FOOD CONSUMPTION
In males of the high-dose group, feed consumption was below control levels at several time points. In females of the high-dose group, this occured once in the main study group, but consumption was almost continuously below control level in the recovery group. At the end of the recovery period, high-dose males consumed slightly more than the control and high-dose females consumed slightly less the the control group, but neither difference was statistically significant.

CLINICAL CHEMISTRY
In high-dose males, bilirubin and triglyceride levels were reduced. Cholesterol was elevated in the mid- and high-dose males and alanine aminotransferase (ALT) only in the high-dose group.
Females of the high-dose group exhibited increased levels of urea nitrogen. Bilirubin and triglyceride levels were below control levels in the mid- and high-dose groups, and cholesterol levels were higher relative to controls.
In the recovery males, ALT, bilirubin and triglycerides were lower than controls for the high-dose group, and cholesterol levels were increased, though not statistically significant. In the recovery females, no effects were observed.

URINALYSIS
Dose-dependent elevations in ketone levels were noted in females, which returned to control levels by the end of the recovery period.

ORGAN WEIGHTS
- Liver weights were increased in high-dose animals. At the mid-dose level, relative liver (to body) weight was increased in males and females and absolute and relative (to brain) liver weights were increased in mid-dose males only. At the end of the recovery period, liver weight differences had largely disappeared.
- Differences in kidney weight (viz. increased relative kidney/body weight in mid- and high-dose females, increased in relative kidney/brain weight in high-dose females, decreased relative kidney/brain weight in high-dose males at the end of the recovery period) reflect differential growth in body and brain weights rather than any significant differrences in kidney weight. This also applies to the observed decrease in adrenal gland weight in mid- and high-dose females, and to the increase in relative testes weight in the high-dose males after the exposure period.
- Absolute brain weight for the high-dose females was decreased at the end of exposure, but not after the recovery period. In the high-dose males, an increased relative brain/body weight was observed at the end of exposure and after the recovery phase. These differences reflect growth retardation early in the animals' lives.

HISTOPATHOLOGY: NON-NEOPLASTIC
Hepatocyte hypertrophy with dose-dependent severitywas observed in all animals of the high-dose group and in males of the mid-dose group. The hypertrophy was characterized by an increase in the cell size with compression of the adjoining sinusoids. Affected hepatocytes were primarily located in the portal area, but in the high-dose group the lesions were diffuse and more eosinophilic. Furthermore, a decreased incidence of small cytoplasmic vacuoles was observed in a dose-dependent manner.
Slight increases in cytoplasmic basophilia of the proximal convoluted tubules were observed in the kidneys of the high-dose group. The same cells contained small numbers of cytoplasmic vacuoles, brush borders were absent and nuclei were slightly enlarged and vesicular with marginated chromatin (4/10 males and females were affected).
Stomach lesions in high-dose males consisted of minimal acanthosis and hyperkeratosis of the non-glandular forestomach (6/10 animals were affected).
At the end of the recovery period, the observed changes were largely reversed. Single animals still showed hepatocyte hypertrophy and decreased severity and amount of vacuolization.

Effect levels

Dose descriptor:

NOAEL

Effect level:

ca. 200 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: Reduced feed consumption and decreased rate of body weight gain

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

The liver enlargement was considered to be primarily an adaptive change rather than a toxic effect.