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Toxicological information

Repeated dose toxicity: oral

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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: Guideline-Studie (GLP, QAU)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1992
Report Date:
1992

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
yes
Remarks:
Only clinical signs, mortality and pathology/histopathology (histotechnical processed and a cytochemical presentation of peroxisomes with diaminobenzidine in the hepatocytes of the liver was performed in semi-thin sections)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): dibutyl phthalate (test substance number 89/449)
- Analytical purity: > 99% (GC)
- Lot/batch No.: P .66 A/89
- Other: positive control substance (name as cited in study report: bis-[2-ethylhexyl] phthalate [test substance number 89/588]; analytical purity: > 99.5% [GC]; lot/batch No.: P .144 A/89, B .63A)

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: male and female Wistar rats (Chbb = THOM, SPF) used were from Dr . Karl Thomae GmbH, Biberach/Riss, FRG, and clinically free from any signs of disease
- Age at study initiation: 35 days at supply
- Housing: during the study period the rats were housed singly in type DK III stainless steel wire mesh cages, supplied by BECKER & CO., Castrop-Rauxel, FRG (floor area about 800 cm2)
- Diet (e.g. ad libitum): the feed used was ground KLIBA rat/mouse/hamster maintenance laboratory diet GLP 343 meal of KLINGENTALMÜHLE AG, CH-4303 Kaiseraugst
- Water (e.g. ad libitum): drinking water
- Acclimation period: 7-day

ENVIRONMENTAL CONDITIONS
The animals were accommodated in a fully air-conditioned room
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- 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:
The test substance preparations were prepared by weighing in the test substance and mixing this thoroughly with a small amount of the feed. This was then mixed in a BOSCH household mixer. An appropriate amount of feed was then added to obtain the desired concentration, and mixing was carried out for about 10 minutes in a GEBR. LÖDIGE laboratory mixer. Different amounts of DIBUTYL PHTHALATE and Bis-(2-ethylhexyl)- phthalate were added, depending on the test groups to the feed at the end of the adaptation period

DIET PREPARATION
- Rate of preparation of diet (frequency): the test substance preparations were prepared in up to 2-week intervals for reasons of the confirmed stability over 32 days.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical results verified the correctness of the test substance concentrations in the diet with two exceptions.
Duration of treatment / exposure:
90 days
Frequency of treatment:
continuously via food
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
ca. 30, 152, 752 mg/kg bw per day
Basis:
other:
Remarks:
Doses / Concentrations:
0, 400, 2000 and 10000 ppm
Basis:
nominal in diet
No. of animals per sex per dose:
3 (see Table 1)
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: the doses were chosen on the basis of the 28-day feeding study with test substance and positive control substance carried out at BIBRA, in which concentrations of 0.02, 0.05, 0.1, 0.5, 1.0 and 2.59 in the diet were used. According to preliminary results, the no effect level of test substance was at 0.1 % and of positive control substance is at 0.05%, which is possibly equivalent to a respective test substance intake of about 100 and 50 mg/kg body weight. Due to the above mentioned findings dose levels for test substance of 400, 2000 and 10000 ppm and for positive control substance 1400 ppm were chosen. The study was carried out from August 29, 1989 (day 0) to December 07, 1989 (last day of perfusion fixation).
- Rationale for selecting satellite groups: no satellite group used
Positive control:
Bis-(2-ethylhexyl)- phthalate in a dosage of 1,400 ppm via the diet over 3 months as a positive control group in respect to the number and morphology of the liver peroxisomes

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: a check for dead and moribund animals was carried out twice a day (Mondays to Fridays) or once a day (Saturdays, Sundays and on public holidays).

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: the animals were inspected for evident signs of toxicity twice a day (Mondays to Fridays) or once a day (Saturdays, Sundays and on public holidays). In addition, the animals were also subjected to an additional exact clinical examination once a week.
Sacrifice and pathology:
At the end of the study the animals were sacrificed by perfusion fixation followed by gross-pathological assessment. From all animals the liver was removed, histotechnical processed and a cytochemical presentation of peroxisomes with diaminobenzidine in the hepatocytes of the liver was performed in semi-thin sections.
Other examinations:
None
Statistics:
None

Results and discussion

Results of examinations

Details on results:
CLINICAL SIGNS AND MORTALITY
- The administration of the test substance and Bis-(2-ethylhexyl)-phthalate as addition to the diet in different dosages did not lead to any disturbances of the general state in any test animal
- During the entire administration period no animal died prematurely.

GROSS PATHOLOGY AND HISTOPATHOLOGY: NON-NEOPLASTIC
At the end of the study the animals were sacrificed by perfusion fixation followed by gross-pathological assessment. From all animals the liver was removed, histotechnical processed and a cytochemical presentation of peroxisomes with diaminobenzidine in the hepatocytes of the liver was performed in semi-thin sections.

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
152 mg/kg bw/day (nominal)
Sex:
male/female
Dose descriptor:
LOAEL
Effect level:
752 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: LOAEL based on peroxisomal proliferation

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion