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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:
disregarded due to major methodological deficiencies
Study period:
July 1980 - December 1980
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: Secondary effects due to irritation. Thus, meets definition "This includes studies or data from the literature/reports in which there are interferences [i.e. irritation] between the measuring system and the test substance".

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1980
Report Date:
1980

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)
Deviations:
yes
Remarks:
Only two dose groups (plus vehicle control) were used; toxicity occured at both doses. The study report indicates that the test substance application site remained uncovered.
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Substance type: Clear, oil-like liquid
- Physical state: Liquid
- Analytical purity: Assumed to be 100% for the purposes of dosing
- Lot/batch No.: ECA-6371
- Expiration date of the lot/batch: July 1981
- Stability under test conditions: No data
- Storage condition of test material: Room temperature

Test animals

Species:
rabbit
Strain:
New Zealand White
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: HARE rabbits for Research, Marland Breeding Farms Inc., 521 Burnet Meadow Road, P.O. Box X, Hewitt, New Jersey, 07421
- Age at study initiation: No data
- Weight at study initiation: Males 1.7–2.8 kg; Females 1.6-2.9 kg.
- Fasting period before study: No data
- Housing: Individually in stainless steel cages.
- Diet: ad libitum
- Water: e.g. ad libitum
- Acclimation period: 16 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 60-78 deg F (
- Humidity (%): 43-75%
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): 12 hours dark / 12 hours light

IN-LIFE DATES: From: 11 August 1980 To: 8 September 1980

Administration / exposure

Type of coverage:
open
Vehicle:
other: Primol 185 (mineral oil)
Details on exposure:
TEST SITE
- Area of exposure: the dorsal surface was clipped from the suprascapular area to the hind quarters, approximately 10cm
- % coverage: No data
- Type of wrap if used: To avoid the possibility of applying test substance to the control dorsal area (untreated), a folded paper towel was held over the left suprascapular dorsal surface of all treated and control animals during the application of the vehicle and test substance
- Time intervals for shavings or clipplings: Rabbits were reclipped on Monday and Thursday of each week throughout the study

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The backs of rabbits were gently wiped with paper towels approximately 6 hours after exposure to remove excessive test substance if necessary.
- Time after start of exposure: Approximately 6 hours.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2mL of test substance in vehicle per kg body weight
- Concentration (if solution): 5 and 25% (based on previous toxicity studies conducted with this substance)
- Constant volume or concentration used: Constant volume used.

VEHICLE
- Justification for use and choice of vehicle (if other than water): No data
- Amount(s) applied (volume or weight with unit): 2mL/kg body weight
- Lot/batch no. (if required): 2938
- Purity: No data

USE OF RESTRAINERS FOR PREVENTING INGESTION: Yes, all rabbits were fitted with Elizabethan collars
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
Duplicate 10mL samples were taken from each weekly batch of dosing solutions prepared and sent to the study Sponsor for analytical confirmation of test substance concentration.
Duration of treatment / exposure:
28 days
Frequency of treatment:
5 days/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
approx. 100mg/kg body weight
Basis:
nominal per unit body weight
Remarks:
Doses / Concentrations:
approx. 500 mg/kg body weight
Basis:
nominal per unit body weight
No. of animals per sex per dose:
10 males and 10 females per dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Based on previous toxicity studies with the test substance
- Rationale for animal assignment (if not random): Random
Positive control:
No

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Observations for mortality and gross signs of toxicologic or pharmacologic effects were performed twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed physical examination for signs of local or systemic toxicity, pharmacologic effects and palpation of tissues masses was performed weekly.

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: Recorded daily, 7 days/week. Observations were made immediately prior to the application of the test substance.

BODY WEIGHT: Yes
- Time schedule for examinations: Once pretest, 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: No data

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 data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Pretest and on Days 27-30.
- Anaesthetic used for blood collection: No
- Animals fasted: Yes
- How many animals: All survivors
- Parameters checked in List 1 were evaluated

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Pretest and on Days 28-30
- Animals fasted: Yes
- How many animals: All survivors
- Parameters checked in List 2 were examined.

URINALYSIS: No data

NEUROBEHAVIOURAL EXAMINATION: No data
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
Complete gross pstmortem examinations were performed on all survivors at termination as well as on all animals either dying spontaneously or killed in extremis during the study. The examinations included examination of the external surface, all orifices, the cranial cavity, carcass, the external and cut surface of the brain and spinal cord, the thoracic, abdominal and pelvic cavities and their viscera, and the cervical tisses and organs.

ORGAN WEIGHTS: Yes (see List 3)

HISTOPATHOLOGY: Yes (see List 4). Control and high-dose animals only, except for testes where low-dose animals also examined.
Statistics:
Body weight, haematology and clinical chemistry parameters, terminal organ and body weights and organ/body weight ratios were analysed. Mean values of all dose groups were compared to control at each time interval.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Dermal irritation:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not specified
Details on results:
CLINICAL SIGNS AND MORTALITY
All low-dose animals (treated with 5% test substance) survived to terminal sacrifice.
In the high-dose group (25% test substance concentration), one male and three females died during the study between Days 17 and 30.

Low- and high-dose males and females exhibited a greater incidence and severity of emaciation than did control animals throughout the period of test substance administration. Thickening of the outer layer of the skin was observed during the last two weeks of the study in low- and high-dose males and females. A greater incidence of lacrimation was observed in the low- and high-dose males (Week 2) and females (Week 2-4) than in the control animals. All animals treated with the test substance also displayed a higher incidence of mucoidal nasal discharge and staining of the anogenital regio compared to controls.

Both low- and high-dose animals generally exhibited marked, dose-related increases in the incidence and severity of erythema, odema, atonia, desquamation, fissuring, eschar formation and exfoliation. Many of the observations were recorded as being moderate to extreme in severity, particularly in the high-dose group.

BODY WEIGHT AND WEIGHT GAIN
Slight to statistically significant, dose-related decreases in body weight were generally exhibited in treated males and females throughout the course of the study.

FOOD CONSUMPTION
No data.

FOOD EFFICIENCY
No data.

WATER CONSUMPTION
No data.

OPHTHALMOSCOPIC EXAMINATION
No data.

HAEMATOLOGY
Low- and high-dose males and females exhibited slight or statistically significantly lower mean haemoglobin, haematocrit and erythrocyte values than did corresponding control animals after four weeks of treatment.

CLINICAL CHEMISTRY
Low- and high-dose males and females exhibited slight or statistically significant, dose-related increases in mean cholesterol values and slight to statistically significant dose-related reductions in albumin. Although these differences were within normal physiological limits, albumin was considered to be borderline low. Total protein and albumin/globulin ratios were normal.

Slight reductions were generally evident in the terminal plasma, erythrocyte and brain cholinesterase values of the low- and high-dose males and females.

URINALYSIS
No data.

NEUROBEHAVIOUR
No data.

ORGAN WEIGHTS
The absolute and relative (organ/body weight ratio) weights of the testes and epididymides of the low- and high-dose males were markedly lower than control weights, showed a dose-related pattern, and were considered to be indicative of a treatment-related effect. high-dose males and low- and high-dose females displayed slight to statistically significant, dose-related increases in mean absolute and relative weights of the adrenals. Slight to statistically significant, dose-related increases in mean absolute and relative kidney weights were also evident in treated males and females.

GROSS PATHOLOGY
Results of the gross postmortem examinations confirmed a higher incidence of exfoliation, fissuring, eschar formation and desquamation in treated animals. The testes of low- and high-dose males were observed to be markedly smaller than those of control males. Other grossly visible morphologic abnormalities observed during the gross postmortem examinations occurred sporadically and were not considered to be treatment-related.

HISTOPATHOLOGY
The results of the microscopic evaluations revealed that the topical administration of the test substance produced irritating skin lesions in both males and females at both the low and high concentrations. In addition, treatment-related testicular effects were evident in the low- and high-dose males. These testicular changes were characterised by morphologic abnormalities in the seminiferous tubules that consisted primarily of aspermatogenesis, diffuse tubular hypoplasia and a reduced mitotic activity.

Effect levels

open allclose all
Dose descriptor:
NOAEL
Basis for effect level:
other: Adverse effects, both systemic and local to the site of administration, were observed at both test substance concentrations (5% and 25%)
Remarks on result:
not determinable
Remarks:
no NOAEL identified
Dose descriptor:
LOAEL
Effect level:
5 other: %
Based on:
test mat.
Sex:
male/female

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Daily application of the test substance (and 5% and 25% concentration) to the clipped skin of rabbits caused local effects at the site of administration rated as moderate to extreme, as well as systemic effects including decreased body weight, emaciation, slight to statistically significant alterations to some haematoligical and clinical chemistry parameters, slight to statistically significant increases in kidney and adrenal weights and, in males, markedly decreased testes weights, which under microscopic assessment revealed morphologic abnormalities in the seminiferous tubules that consisted primarily of aspermatogenesis, diffuse tubular hypoplasia and a reduced mitotic activity. In the high-dose group (25% concentration), one male and three females died during the study. Based on these findings, a no-observed adverse effect level (NOAEL) cannot be identified from this study.
Executive summary:

In a study of repeated dermal toxicity, test substance CAS 28629-66-5 at concentrations of 5% and 25% in mineral oil was applied to the clipped dorsal skin of groups of 10 male and 10 female New Zealand White rabbits, five days per week for four weeks. The dose volume applied was 2mL/kg body weight. A control group of 10 males and 10 females were treated with mineral oil vehicle alone. Animals were observed daily for mortality, gross signs of toxicity and local irritation, and given a more detailed physical examination weekly. Body weight was measured weekly. Blood for haematology and clinical chemistry was taken pretest and at termination. All animals underwent gross pathological examination at necropsy. Selected organs were weighed and a number of tissues examined microscopically.

One high-dose male and three high-dose females died during the study. There were no deaths in the low-dose group. The test substance caused local skin effects at the site of administration ranked from moderate to severe. Clinical signs of toxicity in treated animals included decreased body weight and emaciation, lacrimation, mucoidal nasal discharge and staining of the anogenital region. Haematology revealed slight or statistically significantly lower mean haemoglobin, haematocrit and erythrocyte values in treated animals compared to control animals. Clinical chemistry found slight to statistically significant increases in cholesterol and decreases in albumin. Slight decreases in terminal plasma, erythrocyte and brain cholinesterase was also generally evident in treated animals. At postmortem, gross examination revealed markedly decreased testes size in both low- and high-dose males, and a higher incidence of exfoliation, fissuring, eschar formation and desquamation in all treatment groups. Testes and epididymides of treated males were found to be markedly lower in weight than in control males. Slight to statistically significant, dose-related increases in kidney and adrenal weight were also seen in treated animals. Histological examination showed testicular changes characterised by morphologic abnormalities in the seminiferous tubules that consisted primarily of aspermatogenesis, diffuse tubular hypoplasia and a reduced mitotic activity.

[The study report documents the testing of three separate test substances for 28-day dermal toxicity; however, only the information relating to the study of CAS 28629-66-5 is presented in this Endpoint Study Record.]