Registration Dossier

Administrative data

Endpoint:
short-term repeated dose toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1992
Reliability:
1 (reliable without restriction)

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EPA OPP 82-2 (Repeated Dose Dermal Toxicity -21/28 Days)
GLP compliance:
yes
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rabbit
Strain:
New Zealand White
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Hazleton Research Products Inc., Kalamazoo, Michigan.
- Age at study initiation: 3 months
- Weight at study initiation: 2.03-2.86 kg (males); 2.09-2.89 kg (females)
- Number of animals per group: 5 males and 5 females per group
- Control animals: Yes (5 males and 5 females)
- Housing: the rabbits were individually hooused in a stainless steel cage
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 17 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 1 °C
- Humidity: 72 ± 4%
- Air changes (per hr):
- Photoperiod: 12-hrs light / 12-hrs dark

IN-LIFE DATES: From: 10 June 1991 (receipt date) To: 18-19 July 1991 (sacrifice date)

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST SITE
- Area of exposure: approximately 15 cm wide
- % coverage: 75% group 1 (control group); 5% group 2; 15% group 3; 65% group 4.
- Type of wrap if used: The test site were wrapped with gauze bandaging and secured with non irritating tape.
- Time intervals for shavings or clipplings: The hair was clipped as necessary during the study period to prevent the test article from becoming matted in the hair and to facilitate accurate observations.

REMOVAL OF TEST SUBSTANCE
- Washing: The test site was wiped with disposable toweling
- Time after start of exposure: approximately 6 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): The test article was administered at dosage levels of 0 (control), 100, 300 and 1000 mg/kg (in terms of actual PBO content) using an appropriate size syringe

USE OF RESTRAINERS FOR PREVENTING INGESTION: no. The test substance was applied on the back of the animal and each animal was housed individually in a stainless steel cage.
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
21 days
Frequency of treatment:
5 days per week
Doses / concentrations
Remarks:
Doses / Concentrations:
0 (control), 100, 300, 1000 mg/kg bw/d
Basis:
nominal per unit body weight
No. of animals per sex per dose:
5/male/0; 5/male/100; 5/male/300; 5/male/1000 mg/kg bw/d.
5/female/0; 5/female/100; 5/female/300; 5/female/1000 mg/kg bw/d.
Control animals:
yes
Details on study design:
- Route of exposition selection rationale: A potential route of exposure to humans is dermal. Therefore, this route of administration was employed in this study.
- Dose selection rationale: The doses chosen cover three different situations of dermal exposition: low, medium and high

Examinations

Observations and examinations performed and frequency:
- CLINICAL SIGNS, MORTALITY
Observations were conducted twice daily to assess mortality, morbidity and signs of overt toxicity. A more detailed physical examination was conducted on each animal on a weekly basis. Individual body weight and food consumption were measured weekly.
BODY WEIGHT
Individual body weights were measured weekly.
FOOD CONSUMPTION
Individual food consumption was measured weekly.
WATER CONSUMPTION
Not recorded
OPHTHALMOSCOPIC EXAMINATION
No
HAEMATOLOGY
Clinical laboratory studies were conducted at pre-initiation and termination.
Blood samples were collected from the central ear artery following an overnight fasting period in which animals had free access to water. Hematological parameters examined were leukocyte and erythrocyte count, hemoglobin, hematokrit, MCV, MCH, MCMC, platelets and differential leukocyte count. Clinical parameters measured were sodium, potassium, chloride, calcium, phosphorous, alkaline phosphatase, bilirubin, AST, ALT, urea nitrogen, creatinine, protein, albumin, globulin, cholesterol and glucose.
CLINICAL CHEMISTY
Clinical laboratory studies were conducted at pre-initiation and termination.
Clinical parameters measured were sodium, potassium, chloride, calcium, phosphorous, alkaline phosphatase, bilirubin, AST, ALT, urea nitrogen, creatinine, protein, albumin, globulin, cholesterol and glucose.
URINALYSIS
Not performed.
Sacrifice and pathology:
ORGAN WEIGHTS, GROSS AND HISTOPATHOLOGY
All survivors were euthanized by intravenous sodium pentobarbital followed by exsanguinations.. All animals received a complete postmortem examination under the direct supervision of a pathologist. At necropsy, the adrenals, kidneys, liver and testis were weighed. Microscopic examination of sections preserved in formalin fixed hematoxylin-eosin stained paraffin were performed on all animals. Macroscopic lesions in any other tissue were also examined.
Statistics:
Body weights, food consumption, haematological and biochemical parameters and absolute and relative organ weights were analyzed using analysis of variance (one way classification) and Bartelett’s test for homogeneity of variance. Treatment groups were compared to the control group by sex using the appropriate t-statistic. For determination of significance of differences, Dunnett’s multiple comparison tables or pair wise comparison with a Bonferroni correction were used. All statistical tests are two-tailed, with p ≤ 0.05 and p ≤ 0.01 used as levels of significance.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
No remarkable findings were noted
Dermal irritation:
effects observed, treatment-related
Description (incidence and severity):
See "Details on results"
Mortality:
no mortality observed
Description (incidence):
No remarkable findings were noted
Body weight and weight changes:
no effects observed
Description (incidence and severity):
See "Details on results"
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
See "Details on results"
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
See "Details on results"
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
See "Details on results"
Urinalysis findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
See "Details on results"
Gross pathological findings:
no effects observed
Description (incidence and severity):
See "Details on results"
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
See "Details on results"
Details on results:
CLINICAL SIGNS AND MORTALITY
All test animals survived until termination of the study.

DERMAL IRRITATION
Treatment related effects were limited to minor changes in the skin at the application site.Dermal irritation was observed in all three groups, altough a lesser extent and incidence at 100 mg/kg level.
Very slight erythema and edema appeared by day 5 and generally persisted. At 300 and 1000 mg/kg noted desquamation and fissuring.
At necropsy, findings in the treated animals included red, disclored areas, scabs and thickening of the skin. Microscopic evaluation of the application site skin revealed trace to moderate acanthosis, hyperkeratosis and chronic inflammation of the epidermis. The severity of these lesions increased with increasing dose in both sexes. Acanthosis and chronic inflammation were also noted in the control group, although at a lower incidence and all of the lesions were graded as trace or mild.

BODY WEIGHT AND WEIGHT GAIN
There were no remarkable changes or differences observed in body weights during the study period (see the table reported in "Any other information on results incl. tables")

FOOD CONSUMPTION
Although food consumption was generally lower in treated animals there was no statistically significant difference between controls and treated groups.

WATER CONSUMPTION
No data

OPHTHALMOSCOPIC EXAMINATION
Not done

HAEMATOLOGY
No effects

CLINICAL CHEMISTRY
No effects.

URINALYSIS
Not done

ORGAN WEIGHTS
There were no significant differences in absolute and relative organ weights for either sex among the treatment groups.

GROSS PATHOLOGY AND HISTOPATHOLOGY
No effects

Effect levels

open allclose all
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: does not cause systemic toxicity to the rabbit at a 3 week repeated dermal application of 1000 mg/kg b.w.
Key result
Dose descriptor:
LOAEL
Effect level:
>= 1 000 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: does not cause systemic toxicity to the rabbit at a 3 week repeated dermal application of 1000 mg/kg b.w.

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

The table reported below has been extracted from the full study report N. 542 -007

Dodsage Level

(mg/kg)

 

Mean Body Weigth, grams

(percent difference from control)

Males

Females

0 (Control)

2780

2761

100

2691 (- 3,2)

2640 (- 4,4)

300

2825(+ 1,6)

2868 (+ 3,9)

1000

2777 (- 0,1)

2790 (+ 1,1)

Applicant's summary and conclusion

Conclusions:
MATERIALS AND METHODS
A repeated dose dermal toxicity study according to US EPA Pesticide Assessment Guidelines, Subdivision F, 82-2.
Piperonyl butoxide was administered dermally, undiluted as received, at dosage levels of 100, 300 and 1000 mg/kg b.w. once daily, five days per week for three weeks to five rabbits per sex and dosage group. A control group received mineral oil at the same treatment regime.
Observations were conducted to assess mortality, morbidity and signs of ouvert toxicity. Clinical signs, body weight and food consumption were recorded. In addition haematology and clinical biochemistry, gross pathology and histopathology were investigated.

RESULTS AND DISCUSSION
Treatment related effects were limited to minor changes in the skin at the application site.

CONCLUSION
The dermal administration of the substance up to 1000 mg/kg for 21 days caused no systemic toxicity in male and female rabbits. Topical administration did produce a range of macroscopically and microscopically observed dermal irritation consisting of changes in the epithelium which were dose related in incidence and degree.
Finally, it is concluded that Piperonyl Butoxide does not cause systemic toxicity to the rabbit at a 3 week repeated dermal application of 1000 mg/kg b.w.

LO(A)EL > 1000 mg/kg b.w.
NO(A)EL = 1000 mg/kg b.w.

Reliability: 1
Deficiencies: No