Registration Dossier

Administrative data

Endpoint:
short-term repeated dose toxicity: dermal
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1970
Report Date:
1970

Materials and methods

Principles of method if other than guideline:
o-Phenylenediamine was applied at 300 mg/kg/day to the bare skin of rabbits for six hours a day for ten days over a two week period.
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): o-Phenylenediamine
- Purity: approximately 98.5-99%

Test animals

Species:
rabbit
Strain:
other: Albino
Sex:
male

Administration / exposure

Type of coverage:
occlusive
Vehicle:
other: hydrophilic ointment
Details on exposure:
TEST SITE
- Area of exposure: clipped intact trunk skin
- Type of wrap if used: wrapped with polyvinylidene chloride film, stretch gauze bandage, and elastic tape

REMOVAL OF TEST SUBSTANCE
- Washing: washed with water
- Time after start of exposure: 6 hours

TEST MATERIAL
- Amount applied: 300 mg/kg of body weight of a 10% paste in hydrophilic ointment (ca. 7.6 g).
- Constant volume or concentration used: yes

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
300 mg/kg of body weight of a 10% paste in hydrophilic ointment (ca. 7.6 g). Doses were calculated each day.
Duration of treatment / exposure:
6 hours
Frequency of treatment:
six hours a day for ten days over a two week period
Doses / concentrations
Dose / conc.:
300 other: mg/kg bw
No. of animals per sex per dose:
Six male albino rabbits.
Control animals:
yes

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Not reported

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Not reported

DERMAL IRRITATION (if dermal study): Yes
- Time schedule: Not reported

BODY WEIGHT: Yes
- Time schedule: Not reported

FOOD CONSUMPTION: Yes
- Time schedule: Not reported

WATER CONSUMPTION: Yes
- Time schedule: Not reported

HAEMATOLOGY: Yes
- Time schedule for collection of blood: On the morning of the tenth treatment, blood taken from the ears of three test and three control animals for a Complete Blood count. On the last day, blood was again taken from the ears of the remaining rabbits for a Complete Blood Count.
- Anaesthetic used for blood collection: Not reported
- Animals fasted: No
Sacrifice and pathology:
HISTOPATHOLOGY: After the 10th treatment, half of the group was sacrificed for histopathologic examination and the remaining test and control animals were observed for an additional 14 days before sacrifice.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
On the fourth treatment day the animals appeared to be squinting and were light sensitive. By the fifth day the eyelids were red and swollen with thick yellow discharge. As the test continued, they became swollen, (almost closed), very red with copious thick discharge which in several cases contained blood. The edges of the top conjunctiva appeared blanched in some cases. During the recovery period the swelling and redness subsided considerably but was still evident at the end of the test. This appeared to be an allergic reaction as might be expected from a sensitizer.

The test animals were lethargic but hyper-responsive to room noise from the fourth treatment day and into recovery. Two rabbits had substantial weight loss, reduced appetite, and died during treatment. One was touch sensitive and nervous on the fourth and fifth day and was dead on the sixth day. The other was nervous the second week and shortly after applying the tenth treatment, died with convulsive gasping and cyanosis. By the second week of the recovery period the survivors' behavior seemed normal.

After two treatments small hemorrhagic areas appeared under the skin which became worse during the first week and then disappeared as the injured skin began drying and sloughing off. Skin erythema was moderate to strong during treatment and became progressively better during recovery. The skin appeared grossly normal by the end of recovery. The skin of the control group showed small hemorrhagic areas by the third to fifth treatment day and by the second week the injured skin began drying and sloughing off. The skin erythema was mild to strong during treatment and returned to normal quickly during the recovery period. The control animals showed no other signs.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
During the test period, rabbits lost 11% body weight and returned to original body weight at nine days post treatment.
Description (incidence and severity):
Food and water consumption corresponded with weight loss.
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
Food and water consumption corresponded with weight loss.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
The haemoglobin concentrations (10.5 g/100 mL vs 14.0 g /100 mL control), red blood cell count (5.3 x 10e6/mm3 vs 7.2 x 10e6/mm3 control), and hematocrit (32% vs 42%) were all decreased when measured at the end of treatment. These values were near normal at the end of the recovery period.
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were no significant differences between the test and control organ weights.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
o-Phenylenediamine produced generalized toxemia: exudative hemorrhagic reaction of the skin and acute suppurative conjunctivitis. After recovery for two weeks, no residual effects were seen in the organs and skin. The eyes had begun to heal.

Effect levels

Dose descriptor:
NOAEL
Remarks on result:
not determinable
Remarks:
no NOAEL identified

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Executive summary:

o-Phenylenediamine applied at 300 mg/kg/day to the bare skin of rabbits for six hours a day for ten days over a two week period produced accumulative toxic effects and resulted in two deaths during treatment. The survivors were debilitated, lost body weight and had anemia. The compound was strongly irritating and produced sensitization as evidenced by suppurative conjunctivitis beginning on the fourth day. The surviving rabbits showed a good recovery pattern and no irreversible damage was apparent.