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Toxicological information

Repeated dose toxicity: dermal

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Administrative data

Endpoint:
short-term repeated dose toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
16 June 1983 - 15 July 1983
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-like (QA signature), guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1983
Report Date:
1983

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)
Deviations:
no
GLP compliance:
yes
Remarks:
GLP-like (QA signature)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Batch/Lot Number: 84281
- Physical state: amber viscous liquid
- Analytical purity: 100%
- Stability under test conditions: undetermined
- Storage condition of test material: 60 - 85°F

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Kingston, NY, USA
- Age at study initiation: Approximately 6 weeks old
- Weight at study initiation: Males: 179-210g; Females: 137-161 g
- Fasting period before study: Not applicable
- Housing: Individually in elevated stainless steel cages.
- Diet: Purina Lab Chow #5002 was provided ad libitum. Fresh food was presented weekly
- Water: tap water was made available ad libitum
- Acclimation period: Males: 16 days; Females 17 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): controlled environment (temperature monitored twice daily), but no temparature information provided.
- Humidity (%): controlled environment (humidity monitored twice daily), but no temparature information provided.
- Air changes (per hr): controlled environment (temperature monitored twice saily), but no air change information provided.
- Photoperiod: 12 hour light/dark cycle

IN-LIFE DATES: From: To: 16/06/1983-15/07/1983

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST SITE
- Area of exposure: dorsal surface and sides
- % coverage: approximately 10 % of the body surface
- Type of wrap if used: porous gauze covered with an elastic adhesive bandage
- Time intervals for shavings or clipplings: the animals were reclipped weekly

REMOVAL OF TEST SUBSTANCE
- Washing (if done): the skin was wiped after exposure but not washed
- Time after start of exposure: 6 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): An appropriate amount of test material was applied based on the most recent weekly bodyweight
- Concentration (if solution): Not applicable
- Constant volume or concentration used: yes
- For solids, paste formed: Not applicable

VEHICLE
- Justification for use and choice of vehicle (if other than water): Not applicable
- Amount(s) applied (volume or weight with unit): Not applicable
- Concentration (if solution): Not applicable
- Lot/batch no. (if required): Not applicable
- Purity: Not applicable

USE OF RESTRAINERS FOR PREVENTING INGESTION: No
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
The test material was not diluted with a vehicle. It was applied as received at the intended dose (amount of test material).
Duration of treatment / exposure:
4 weeks
Frequency of treatment:
5 days per week
Doses / concentrations
Remarks:
Doses / Concentrations:
100, 300 or 1000 mg/kg bw/day (m/f)
Basis:
nominal per unit body weight
No. of animals per sex per dose:
5/sex/dose
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: No information provided
- Rationale for animal assignment: animals were assigned to groups randomly
- Rationale for selecting satellite groups: Not applicable
- Post-exposure recovery period in satellite groups: Not applicable
Positive control:
Not applicable

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily, 7 days/week
- Cage side observations checked in table No. 1 were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily, 5 days/week prior to each application of test material

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: daily, 5 days/week just prior to the next application of test material

BODY WEIGHT: Yes
- Time schedule for examinations: Twice pre-test, weekly during treatment and terminally (after fasting)

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No
- Time schedule for examinations: Not applicable

OPHTHALMOSCOPIC EXAMINATION: No
- Time schedule for examinations: Not applicable
- Dose groups that were examined: Not applicable

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At termination
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes, animals were fasted overnight prior to blood collections
- How many animals: all animals were tested
- Parameters checked in table No. 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At termination
- Animals fasted: Yes, animals were fasted overnight prior to blood collections
- How many animals: all animals were tested
- Parameters checked in table No. 1 were examined.

URINALYSIS: No
- Time schedule for collection of urine:
- Metabolism cages used for collection of urine: Not applicable
- Animals fasted: Not applicable

NEUROBEHAVIOURAL EXAMINATION: No
- Time schedule for examinations: Not applicable
- Dose groups that were examined: Not applicable
- Battery of functions tested: Not applicable

Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table 1)
HISTOPATHOLOGY: Yes (see table 1)
Other examinations:
The following organs were weighed: adrenals, liver, kidneys, testes.
Statistics:
Body weight, food consumption, hematology and clinical chemistry parameters, organ weights and organ/body weight ratios were analysed. Mean values of all dose groups were compared to control at each time interval.
Statistical evaluation of equality of means was made by the appropriate one way analysis of variance technique, followed by a multiple comparison procedure if needed. First, Bartlett's test was performed ot determine if groups had equal variance. If the variances were equal, parametric procedures were used (one way ANOVA using the F distribution to assess significance). If significant differences among the means were indicated, Dunnett's test was used to determine which means were significantly different from the control. If a nonparametric proceduse for testing equality of means was needed, the Kruskal-Wallis test was used, and if differences were indicated a summed rank test (Dunn) was used to determine which treatments differed from control.
A statistical test for trend in the dose levels was also performed. In the parametric case (i.e. equal variance) standard regression techniques with a test for trend and lack of fit were used. In the nonparametric case Jonckheere's test for monotonic trend was used.
The test for equal variance (Bartlett's) was conducted at the 1 %, two-sided risk level. All other statistical tests were conducted at the 5 % and 1 %, two-sided risk level.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Dermal irritation:
effects observed, treatment-related
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY

All animals survived throughout the study. One control male exhibited occasional abnormalities (dry red ocular or nasal discharge, unthrifty coat). No evidence of systemic abnormalities was seen in any of the treated animals.

Evidence of mild dermal irritation (erythema and/or desquamation at the dose site) was noted in some animals (please refer to table 2 below). Erythema was generally noted near the end of the first week of dosing and occurred in individual animals on one to three occasions. Desquamation occurred primarily during the third and fourth weeks. no evidence of severe dermal irritation or necrosis was seen in any animal.

BODY WEIGHT AND WEIGHT GAIN

All animals gained weight over the course of the study. Mean values for control and treated animals were considered comparable.

FOOD CONSUMPTION

Food consumption values for control and treated groups were considered comparable.

FOOD EFFICIENCY

Not applicable

WATER CONSUMPTION

Not applicable

OPHTHALMOSCOPIC EXAMINATION

Not applicable

HAEMATOLOGY

Evaluation of hemoglobin, hematocrit, total erythrocyte, platelet and total and differential leukocyte values revealed no evidence of an effect of administration of the test material on these parameters.

CLINICAL CHEMISTRY

Clinical chemistry values for control and treated groups were considered comparable. There was no evidence of an effect of test material administration.

URINALYSIS

Not applicable

NEUROBEHAVIOUR

Not applicable

ORGAN WEIGHTS

Weights of adrenals, kidneys, liver and testes were comparable among groups as were organ/body weiight ratios for these organs.

GROSS PATHOLOGY

The effects noted are summarised in table 3 below. There were no effects that were considered to be related to test material administation.

HISTOPATHOLOGY: NON-NEOPLASTIC

Microscopic examination of liver and kidney sections from control and high-dose animals revealed no changes considered to be related to test material administration.

Minimal to slight epidermal hyperplasia was noted in the treated sin of 6 of 9 animals form the high dose group (3/4 males, 3/5 females). similar changes were not seen in skin from control animals.

HISTOPATHOLOGY: NON-NEOPLASTIC

Not applicable

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Systemic effects
Dose descriptor:
LOAEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Local effects at application site (mild skin irritation)

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Table 2: No. of animals exhibiting dermal irritation on one or more occasion

 

Males

 

Females

 

Dose group (mg/kg bw/day)

Erythema

Desquamation

Erythema

Desquamation

0

0/5

0/5

0/5

0/5

100

0/5

0/5

1/5

1/5

300

1/5

0/5

3/5

1/5

1000

4/5

1/5

5/5

3/5

 

Table 3: Organs examined and effects noted at necropsy

 

No. of effects seen/No. of animals examined

Male

Female

Dose group (mg/kg bw/day)

0

100

300

1000

0

100

300

1000

Organ and effects

 

 

 

 

 

 

 

 

Liver

  Discolored

  Surface: irregularities

 

0/5

0/5

 

1/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

1/5

0/5

Kidneys

  Pelvis: Dilated

 

1/5

 

0/5

 

1/5

 

1/5

 

0/5

 

0/5

 

1/5

 

0/5

Treated skin

0/5

0/5

0/5

0/5

0/5

0/5

0/5

0/5

Untreated skin

 Discolored

 

1/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

Thymus

 Discolored

 

0/5

 

1/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

Tail

 Discolored

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

Lungs

 Discolored

 

0/5

 

0/5

 

1/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

Incisors

 Maloccluded

 

1/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

Lymph node

 Discolored

 Enlarged

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

 

0/5

0/5

Ureters

 Large

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

0/5

 

1/5

 

0/5

Ovaries

 Cysts

 

0/0

 

0/0

 

0/0

 

0/0

 

0/5

 

0/5

 

0/5

 

0/5

Adrenal glands

0/5

0/5

0/5

0/5

0/5

0/5

0/5

0/5

Applicant's summary and conclusion

Conclusions:
Administration of 100, 300 or 1000 mg/kg/day of the test substance, five days/week for four weeks to the clipped dorsal surface of rats did not cause any systemic effect. Test substance-related effects were limited to local, portal of entry, mild dermal irritation. The no observed effect level for systemic toxicity was 1000 mg/kg/day; the lowest observed adverse effect level for local, portal of entry effects (mild skin irritation; 1/5 females) was 100 mg/kg/day. In accordance with EU CLP Regulation (EC) No. 1272/2008 classification of this substance for repeat dose toxicity via the dermal route is not required.
Executive summary:

Test Guidance

Twenty-eight day repeat dose dermal toxicity study performed according to OECD TG 410.

Method and Material

The test material was applied to the shaved dorsal area and sides of rats (5 animals/sex/dose) at dose levels of 0, 100, 300 or 1000 mg/kg bw/day, five days/week for four weeks. After application of the test material the dose site was covered with a semi-occlusive dressing for 6 hours, after which the dressing was removed and the skin wiped free of any excess test material. The effect of the test material on the rats was evaluated according to physical appearance, dermal irritation, body weight, food consumption, haematology, clinical chemistry, organ weights, gross and microscopic pathology.

Results

There were no mortalities and no evidence of systemic toxicity was seen in any of the groups during the study. Evidence of mild dermal irritation (erythema after the first week of dosing and/or desquamation during the third and fourth week) was seen in two females at 100 mg/kg bw/day, one male and most females at 300 mg/kg bw/day and most animals at 1000 mg/kg bw/day. No severe dermal irritation or evidence of deep tissue damage was apparent, however. Microscopic examination of treated skin from control and high dose animals revealed minimal to slight epidermal hyperplasia in several (6 of 9) animals in the 1000 mg/kg bw/day group. Evaluations of body weights, food consumption, clinical chemistry studies, organ weights, organ/body weight ratios and microscopic examination of liver and kidneys from animals in the 1000 mg/kg bw/day group did not reveal any effects considered to be related to administration of the test material. The NOEL for systemic toxicity was determined to be 1000 mg/kg bw/day. The LOAEL for local effects was 100 mg/kg bw/day based on mild skin irritation observed in 1/5 females.

Conclusion

In accordance with EU CLP Regulation (EC) No. 1272/2008 classification of this substance for repeat dose toxicity via the dermal route is not required.