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Toxicological information

Repeated dose toxicity: oral

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Administrative data

sub-chronic toxicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Non-GLP near-guideline study, acceptable for assessment, please refer to IUCLID section 13 for read across justification.

Data source

Reference Type:
Thirteen-week toxicity study of d-alpha-tocopheryl acetate (Vitamin E) in Fischer 344 rats.
Abdo KM, Rao G, Montgomery A
Bibliographic source:
Fd Chem Toxic. 24 (10/11): 1043-1050.

Materials and methods

Test guideline
equivalent or similar to guideline
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
GLP compliance:
not specified
Limit test:

Test material

Constituent 1
Reference substance name:
α-tocopheryl acetate
EC Number:
EC Name:
α-tocopheryl acetate
Cas Number:
Molecular formula:
2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-3,4-dihydro-2H-chromen-6-yl acetate
Constituent 2
Reference substance name:
d-alpha-tocopheryl acetate
d-alpha-tocopheryl acetate
Details on test material:
Vitamin E acetate (d-alpha-tocopheryl acetate); according to the authors, purity was >99%

Test animals

Fischer 344
Details on test animals or test system and environmental conditions:
- Source: Charles River Breeding Laboratory, Kingston, NY
- Age at study initiation: 25 days
- Housing: stainless steel cages
- Diet: ad libitum, modified AIN-76 diet
- Water: ad libitum
- Acclimation period: 11 days

Administration / exposure

Route of administration:
oral: gavage
corn oil
Details on oral exposure:
The vitamin (>99% pure and obtained from Eastman Kodak Company, Rochester, NY) was administered in corn oil by gavage at the rate of 3.5 ml/kg
body weight daily for up to 13 weeks.
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
Doses / concentrations
Doses / Concentrations:
125, 500, 2000 mg/kg bw/d

No. of animals per sex per dose:
Control animals:
Details on study design:
Two control groups (10 rats/sex/group) were either administered corn oil by gavage (vehicle control group) or remained untreated (untreated control group).
Post-exposure period: none
Positive control:


Observations and examinations performed and frequency:
Individual blood samples (with or without heparin) for haematology and clinical chemistry determinations were collected by terminally bleeding ten males and ten females from each treatment group via the vena cava at days 5, 45 and 90 of dosing treatment.
Haematology determinations, which included mean corpuscular cell volume, haemoglobin, haematocrit, erythrocyte count and total and differential leucocyte counts, were made on a Baker 7000 cell counter (Baker Instruments, Allentown, PA) . Prothrombin time (PT) and activated partial thromboplastin time (APTT) were determined according to the Coag-a Mate operator's manual (General Diagnostics, Division of Warner-Lambert Co, Morris Plains, NJ) . Fibrinogen levels were determined according to the turbidimetric procedure of Exner using the Cetrifichem 400 (Exner, 1979). Serum samples were analysed for chloride, alkaline phosphatase, thyroxine (T4) and γ-glutamyl transpeptidase (GGT) using the Centrifichem 400 according to the procedures specified by the manufacturer (Baker Instruments, Allentown, PA). Triiodothyronine (T3) and thyroid-stimulating hormone (TSH) were determined by radioimmunoassay procedures (Kieffer, 1974) .
Sacrifice and pathology:
At the end of the treatment period, surviving animals were sacrificed and subjected to complete necropsy.
Organ weights and organ-to-body weight ratios were determined for brain, heart, kidneys, liver, spleen, thyroid and parathyroid (combined), thymus, pituitary gland, ovary, uterus, prostate, epididymis and testes.
In addition to these organs, the spinal cord, eyes, salivary gland, mammary gland, oesophagus, lungs with mainstem bronchi, trachea, stomach, small and large intestine, adrenals, pancreas, urinary bladder, mesenteric lymph node, bone marrow, sternum and sciatic nerve with skeletal muscle were
collected, fixed in 10% buffered formalin, embedded in paraffin, sectioned at 5 ,um and stained with haematoxylin and eosin. All tissues from the control and the high-dose groups were examined microscopically. For the lower-dose groups, only tissues that were designated as targets for vitamin E toxicity were examined (not further specified).
The significance of differences between Vitamin E-treated and control groups was assessed using the Dunnett's multiple comparison procedure. Dose-response trend was assessed by Jonckheere's test. Results were considered significant when the calculated P value was 0.05 or less.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not specified
Details on results:
Seven out of 10 high dose males died during weeks 9-11.
The test substance had no effect on food consumption or body weights.
The relative liver weight was significantly increased in high dose females.
Administration of 2000 mg/kg bw/d caused hematological changes: prolongation of both prothrombin and activated partial thromboblastin (APTT) times, reticulocytosis and a decrease in hematocrit values and hemoglobin concentrations was observed in males; APTT times were also increased in females. Hemorrhagic diathesis was observed in males and females of the high dose group; and increased medullary erythropoiesis was seen in the spleen of one high dose male.
A compound-related increase was also observed in the plasma levels of thyroid-stimulating hormone. However, without a corresponding increase in T3 and T4, and no evidence for increased energy metabolism ( food consumption and body weights were comparable to control groups), the toxcicological relevance was doubted.

The test substance at all dose levels tested caused interstitial inflammation and adenomatous hyperplasia of the lung. The lung lesions were observed in all vitamin E-treated groups, and the incidence and severity increased in a dose-dependent manner. These lesions were characterized by increased cellularity, vascular congestion, thickened alveolar walls and the presence of foamy macrophages (some of which had undergone cell death and degeneration) in the alveolar spaces. A lipid-like yellow pigmentation was often present within either the macrophages or alveoli.
None of these lesions, however, were considered to reflect systemic toxic effects of the test or reference articles. The changes in the lungs were considered to be a local reaction to the aspiration of a-tocopheryl acetate into the lungs that would not occur under conditions of normal use.

Therefore, for the NOAEL derivation the effects in the lungs were not considered.
Because at 500 mg/kg only the APTT value is increased, but not the PT and fibrinogen values, the NOAEL is set at 500 mg/kg.

Effect levels

Dose descriptor:
Effect level:
500 mg/kg bw/day (nominal)
Basis for effect level:
other: see 'Remark'

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion