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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1993-10-25 to 1993-11-09 (In-life)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1996
Report Date:
1996

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Version / remarks:
May 12, 1981
Qualifier:
according to
Guideline:
other: EC Commission directive 92/69/EEC; July 31, 1992
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2,3,6-trimethylphenol
- Physical state: solid
- Analytical purity: 99.7%
- Impurities (identity and concentrations): no data
- Purity test date: no data
- Lot/batch No.: from continuous production "Sumpf K 304", 1990-11-16
- Expiration date of the lot/batch: no data
- Stability under test conditions: yes, confirmed by analysis
- Storage condition of test material: room temperature

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Dr. Karl Thomae GmbH, Biberach/Riss, Germany
- Age at study initiation: 35 days (arrival); 42 days (start of dosing)
- Weight at study initiation: males: 204 (191-223)g; females: 155 (142-164) g
- Fasting period before study: no data
- Housing: individually in mesh wire cages
- Diet: ground Kliba maintenance diet rat/mouse/hamster, 343 meal, supplied by Klingentalmühle AG, Kaiseraugst, Switzerland; ad libitum
- Water: ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: October 04, 1993 (arrival of the animals) To: November 09, 1993 (necropsy)

Administration / exposure

Route of administration:
oral: feed
Vehicle:
acetone
Details on oral exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with (Type of food): standard diet
- Storage temperature of food: no data

VEHICLE
- Justification for use and choice of vehicle (if other than water): no data
- Concentration in vehicle: no data
- Purity: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability of the test substance in the diet was determined analytically before the start of the study.
The homogeneous distribution of the test substance in the diet was proven at the start of the administration period in samples of the high and low concentration. These homogeneity analyses served also as concentration controls. Additionally, concentrational control analyses were performed in samples of the mid concentration at the beginning of the administration period.
Duration of treatment / exposure:
28 days
Frequency of treatment:
continuously in the diet
Doses / concentrations
Remarks:
Doses / Concentrations:
ca. 55, 273, 1346 mg/kg bw/d (600, 3000, 15000 ppm in the diet)
Basis:
actual ingested
No. of animals per sex per dose:
5 rats/sex/group
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale:
In a palatability study, the test substance was administered to groups of 3 male and 3 female Wistar rats at dietary concentrations of 0; 5000 and 15000 ppm for 2 weeks. Following substance-related findings were obtained:
- slight impairment of food consumption at both concentrations
- slight impaired of body weight at both concentrations: the values on day 14 were about 6%/7% (males/females) below control at the high concentration and about 5%/6% below control at the low concentration.
Therefore, the following concentrations were chosen for the present study:
15000 ppm: as highest concentration, corresponding to a test substance intake of at least 1000 mg/kg body weight
3000 ppm: as intermediate concentration
600 ppm: as expected "no observed adverse effect level "
- Rationale for animal assignment: randomly; identification by ear tattoo number
- Post-exposure recovery period: none
Positive control:
none

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): weekly
Food consumption was determined weekly over a period of 7 days and calculated as mean food consumption in grams per animal and day .
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

FOOD EFFICIENCY: No

WATER CONSUMPTION: determined weekly

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: no data
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: 5/sex/group
- Parameters examined:
leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count, prothrombin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: no data
- Animals fasted: No
- How many animals: 5/sex/group
- Parameters examined:
alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, serum-gamma-glutamyltransferase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, magnesium

URINALYSIS: Yes
- Time schedule for collection of urine: no data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters examined:
volume, color, turbidity, nitrite, pH, protein, glucose, ketones, urobilinogen, bilirubin, blood, specific gravity, sediment

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Statistics:
The data were evaluated statistically on the computer systems of the Department of Toxicology of BASF Aktiengesellschaft

Means and standard deviations were calculated for variables.
Methods used (depending on the variable): KRUSKAL-WALLIS test, MANN-WHITNEY U-test.
Significance labels: p<0.05, p> 0.02, p<0.01, p<0.002; depending on method and variable.

Remark: Due to the limited maximum size of this field, no more details on statistics are given herein.

Results and discussion

Results of examinations

Details on results:
CLINICAL SIGNS AND MORTALITY
no substance-related changes

BODY WEIGHT AND WEIGHT GAIN
substance-related changes in high-dose males and females; see below

FOOD CONSUMPTION
substance-related changes in high-dose males and females; see below

WATER CONSUMPTION
no substance-related changes

HAEMATOLOGY
substance-related changes in high-dose males and females and in mid dose females; see below

CLINICAL CHEMISTRY
no substance-related changes

URINALYSIS
no substance-related changes

ORGAN WEIGHTS
substance-related changes in high-dose males and females; see below

GROSS PATHOLOGY
no substance-related changes

HISTOPATHOLOGY: NON-NEOPLASTIC
substance-related changes in high-dose males and females; see below

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
no substance-related changes

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
ca. 273 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
body weight and weight gain
haematology
Dose descriptor:
NOAEL
Effect level:
ca. 55 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
body weight and weight gain
haematology
Dose descriptor:
LOAEL
Effect level:
ca. 1 346 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: highest dose tested
Dose descriptor:
LOAEL
Effect level:
ca. 273 mg/kg bw/day (actual dose received)
Sex:
female
Basis for effect level:
other: Decrease in hematocrit in females

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

According to the authors, the following significant, substance-related findings were obtained in the respective dose groups:

15000 ppm (about 1,346 mg/kg body weight):

  • slightly decreased food consumption in males and females, especially during the first week of the study
  • impairment of body weight in males and females; the values on day 28 were about 4% and 12% below controls, respectively
  • impairment of body weight change in males and females ; the values on day 28 were about 9% and 41% below controls, respectively
  • decrease in red blood cells, hemoglobin and hematocrit in both sexes
  • decrease in mean corpuscular hemoglobin concentration in females
  • increase in polychromasia and cholesterol in both sexes
  • increase in mean corpuscular volume and mean corpuscular hemoglobin in females
  • increase of absolute spleen weight in females and of relative spleen weights in males and females
  • increase of extramedullary erythropoiesis and hemosiderosis of the spleen in males and females
  • slight increase of erythropoiesis in the bone marrow of two males and two females

3000 ppm (about 273 mg/kg body weight):

  • decrease in hematocrit in females

600 ppm (about 55 mg/kg body weight) :

  • no substance-related changes

Thus, substance-related effects were seen at 15000 ppm in both sexes and at 3000 ppm in females. Target organs were spleen, bone marrow and erythron. The only toxic effects observed were signs of marked anemia at the high dose (> 1000 mg/kg body weight) with subsequent enhancement of hematopoiesis in bone marrow and spleen. Only a marginal sign of this effect was seen at the mid dose in females.

The no observed adverse effect level (NOAEL) under the conditions of this study was 3000 ppm (about 273 mg/kg body weight) in males and 600 ppm (about 55 mg/kg body weight) in females.

Applicant's summary and conclusion

Executive summary:

The test substance was administered to rats for 4 weeks at concentrations of 600, 3000 and 15000 ppm for 4 weeks. The concentrations used provided dose levels of approximately 55, 273, and 1346 mg/kg bw/d, respectively. Control animals were administered plain diet.

Administration of the test substance at 15000 ppm adversely affected food consumption, body weight, body weight gain, hematological parameters, spleen weight and induced histological changes in the spleen and bone marrow of males and females. The mid dose level affected a single hematological parameter in females only. No adverse effects were seen in the rats administered 600 ppm.

Under the conditions of this study, the NOAEL was 3000 ppm in the diet (ca. 273 mg/kg bw/d) for males and 600 ppm in the diet (ca. 55 mg/kg bw/d) for females.