Registration Dossier

Toxicological information

Basic toxicokinetics

Currently viewing:

Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
Year of publication: 2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: comparable to guideline study

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2001
Report Date:
2001

Materials and methods

Objective of study:
other: bioavailability of cleavage products
Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: OECD Guideline 412 (Repeated Dose Inhalation Toxicity)
Deviations:
yes
Remarks:
e.g. no data on haematology, because the main subject of the study was the investigation of the bioavailability and not the investigation of toxicity after repeated exposure
GLP compliance:
yes
Remarks:
Performance and documentation was on the basis of GLP according to the German Chemikaliengesetz, but the test was not under supervision of the quality assurance unit

Test material

Reference
Name:
Unnamed
Type:
Constituent
Radiolabelling:
no

Test animals

Species:
rat
Strain:
other: Wistar Crl:(WI)BR
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, Germany
- Age at study initiation: about 3 month
- Weight at study initiation: about 430 g
- Fasting period before study: no data
- Housing: Makrolon cages
- Individual metabolism cages: yes
- Diet: Altromin Pellet-Feed (1324N) or Altromin no Pellet-Feed (1321N) ad libitum
- Water: drinking water ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 40-70%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 h/12 h

Administration / exposure

Route of administration:
other: intratracheal instillation
Vehicle:
other: suspension in isotonic saline containing 1% (w/v) Tween 80
Details on exposure:
- intratrachel instillation of 0.3 ml of suspension

Duration and frequency of treatment / exposure:
once per week for 4 consecutive weeks (day 1, 8, 15, 22 and 29 of the experiment)

Doses / concentrations
Remarks:
Doses / Concentrations:
0, 10, 20 mg/animal per application

No. of animals per sex per dose:
12 males per group in the control and test substance group (6 of the animals were sacrificed on day 31 and 6 animals survived till the end of the 4 weeks post exposure period)
- 6 males in the positive control group
Control animals:
yes, concurrent vehicle
Positive control:
5 times 7.31 mg 3,3'-dichlorobenzidine (CAS 91-94-1) or 5 times 5,6 mg Direct Red 46 (CAS 6548-29-4) per animal

Details on study design:
- Dose selection rationale: ascertainment of the maximum tolerable dose in a pretest


Details on dosing and sampling:
BIOAVAILABILITY STUDY
- Tissues and body fluids sampled: urine, faeces, blood, organs (lung, liver, spleen, kidney, urinary bladder, gastro intestinal tract, mamma)
- Time and frequency of sampling of urine and faeces: on day 1, 2, 3, 7, 8, 9, 14, 15, 16, 21, 22, 23, 28, 29, 30 for the animals which were sacrificed on day 31
- Time and frequency of sampling of urine and faeces: on day 28, 29, 30, 35, 42, 49, 56 for the animals which were sacrificed on day 57 at the end of the post exposure period
- Time and frequency of sampling of blood and organs: after sacrifice on day 31 or 57
Statistics:
no data

Results and discussion

Any other information on results incl. tables

One animal of the negative control group had to be killed in a moribund condition on day 4. There was no direct correlation to the treatment obvious.

Body-weight development was normal.

The test substance was deposited in the lung: The test substance was detectable in macrophages, in the alveolar and intrabronchiolar region. Lung weights were elevated at the end of the exposure period and still elevated at the end of the post exposure period. Bronchio-alveolar hyperplasia and interstitial fibrosis were detected and persisted till the end of the observation period. Pigment loaded macrophages were also detected in lung associated lymph nodes, which showed increased weight and reactive lymphoide hyperplasia. Effects persisted till the end of the observation time.

3,3'-Dichlorobenzidine was not detectable in urine, faeces or bound to haemoglobin after acid hydrolysis of the matrices.

3,3'-Dichlorobenzidine was traceable in the urine of several animals of the control group (2 animals sacrificed on day 31 and 5 animals sacrificed on day 57). The authors of the report concluded that this was probably due to a contamination of the collecting containers.

Pigment content of the lungs determined at the end of the exposure period was about 50 - 60% of the applied dose. The pigment content of the lungs did not decline during the post exposure period.

3,3'-dichlorobenzidine treatment:

Animals treated with 3,3'-dichlorobenzidine excreted about 3% of the applied dose via urine (about 194000 ng on day 1 and 226000 ng on day 29) during the first 24 h after dosing. Urinary excretion declined rapidly thereafter: about 10400 ng/24 h during the second and 1030 ng/24 h during the third 24 hours after dosing. The average urinary excretion on the day before the 5th application was 59 ng/24 h. Excretion of 3,3'-dichlorobenzidine via faeces was delayed in comparison to excretion via urine: About 3.6% ( 262 µg; day 1 and 2) or 2.6% (190 µg, day 29 and 30) were excreted within 48 hours after application.

At sacrifice about 1496 ng 3,3'-dichlorobenzidine were bound per g haemoglobin.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): other: the test substance has a high potential to persist in the lung due to the significantly limited clearance capacity of the lung as a consequence of the particle overload; no 3,3'-dichlorobenzidine is bioavailable from the test substance
Analysis of urine, faeces and haemoglobin revealed that no 3,3'-dichlorobenzidine is liberated from the test substance after intratracheal instillation.
Executive summary:

Male Wistar rats were exposed for five times to 10 or 20 mg test item/animal by intratracheal instillation at weekly intervals. Urine and faeces were collected during the exposure and 28 days observation period, blood was collected at the end of the exposure and post observation period. Urine, faeces and haemoglobin were analysed for the presence of 3,3'-dichlorobenzidine after acid hydrolysis, the pigment content of the lungs was determined gravimetrically and the lungs and lung associated lymph nodes were investigated histopathologically. The pigment was deposited in the lung and persisted in the lung during the observation period. No 3,3'-dichlorobenzidine was detectable in the urine, faeces and haemoglobin of treated rats during the exposure period. Traces of 3,3'-dichlorobenzidine were detected in control animals, probably due to impurities of the analytical instruments.