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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1991
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EPA OPP 83-3 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Portage, Michigan, USA
- Age at study initiation: Approximately: Females 70 days, Males 77 days
- Weight at study initiation: 250-300 g (males) 175-200 (females)
- Housing: Stainless steel wire mesh cage. Deotized Animal Cage Board was placed under each cage and changed regularly.
- Diet: ad libitum. Certified Rodent Chow (5002; Ralston Purina Company, St. Louis, MO)
- Water: ad libitum (tap water available by an autoamtic watering system)
- Acclimation period: approximately 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 19 - 25 °C
- Humidity: 40 - 70 %
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: December 24, 1990 (males birth date); December 31, 1991 (females birth date) To: April 2 – 4, 1991 (maternal sacrifice)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

Dams were dosed daily with undiluted piperonyl butoxiude (PBO) or deionized water on gestation days (gd) 6 through 15.
Dosing substances were administered by gavage using a 16-gauge stainless steel dosing tube, 3.0 inches long attached to a 1000 µl glass syringe.
Details on mating procedure:
- Impregnation procedure: virgin female rats were mated to virgin male rats on a one male to one female basis in stainless steel wire mesh cages (22.5 x 31.0 x 18.0 cm high)
- M/F ratio per cage: 1/1
- Length of cohabitation: The matting period was March 11-14, 1991. gd 0 was March 12-14, 1991. Successful mated female were housed individually for the duration of the study
- Further matings after two unsuccessful attempts: no. Each male was paired only once in this study.
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: Females were checked once daily in the morning for vaginal copulation plugs and the paper board beneath the cages was checked twice daily for dropped copulations plugs. The day of copulation plug was observed was designed gd 0.
Duration of treatment / exposure:
Dams were dosed daily with undiluted PBO or deionized water on gd 6 through 15.
The treatment period was March 18 through March 29, 1991.
Frequency of treatment:
daily
Duration of test:
21 days
Doses / concentrations
Remarks:
Doses / Concentrations:
0 (control), 200, 500, 1000
Basis:

No. of animals per sex per dose:
25/females/0; 25/females/200; 25/females/500; 25/females/1000.
Control animals:
yes
Details on study design:
The doses volumes were based on the dam’s body weight on gestation day 6, the % active ingredient, and the specific gravity of the test substance.
The doses employed were 0, 200, 500 and 1000 mg/kg/day.
These dose levels were selected on the bases of findings from a range-finding study which was conducted at dosage levels of 0, 250, 500, 1000 and 4000 mg/kg/day. In the range-finding study, four of five dams in the 1000 mg/kg/day group and five of five dams in the 4000 mg/kg/day group dosage level died or were sacrificed moribund.

- Rationale for animal assignment: random

Examinations

Maternal examinations:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: All dams were thoroughly examined daily for clinical signs of toxicity (twice daily during the dosing period). In addition, animals were observed twice daily for morbidity and mortality. Maternal body weights were taken on gd 0, 6, 9, 12,15, 18, 21. (See the table "CLINICAL OBSERVATIONS " attached in background material).

BODY WEIGHT: Yes
- Time schedule for examinations: Maternal body weights were taken on gd 0, 6, 9, 12,15, 18, 21 and, the maternal body weights changes were taken on gd 0, 6, 6-9, 9-12, 12-15, 15-18, 18-21. Maternal body weights appeared to be reduced on gd 9 and 12 at 500 and 1000 mg/kg/day. Body weight gain decreases were observed in all treated group for gd 6 to 9. Body weight gains were also reduced for gd 12-15, 15-18 and 6.15 (the entire treatment period) at 500 and 1000 mg/kg/day. Weight gains for gd 6 to 15 were slightly reduced at 500 mg/kg/day mg/kg/day. (See the table "BODY WEIGHT " attached in background material).

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes (no feeding study)
- Maternal food consumption was measured at three-day intervals throughout gestation, gd 0-21 (gd 0-3, 3-6, 6-9, 9-12, 12-15, 15-18, 18-21). The food consumption was slightly depressed at 500 & 1000 mg/kg/day (See the table "FOOD CONSUMPTION " attached in background material).

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21.
- Organs examined: the dams were evaluated for body weight, liver and gravid uterine weight, number of corpora lutea and number and status of implantation sites (including early and later resorptions, dead fetuses, and live fetuses). (See the tables "ORGAN WEIGHTS " and "GESTATIONAL PARAMETERS" attached in background material).
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes.
Examinations included:
- Gravid uterus weight: Yes
- Liver weight: yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
(See the tables "ORGAN WEIGHTS" & "GESTATIONAL PARAMETERS" attached in background material)
Fetal examinations:
- All live fetuses were dissected from the uterus, counted, sexed, weighed, examined for external malformations (including clef palate) and variations. Approximately one-half of the live fetusesw in each litter were examined for visceral malformations. These foetuses were decapitated and the heads were examined for soft tissue craniofacial malformations and variations. The remaining half of the foetuses (intact) in each litter were eviscerated fixed in alcohol, stained with alizarin red S and examined for skeletal malformations and variations.

- There were no treatment-related effects on fetal body weights.
- There were statistically significant differences in individual external, visceral or skeletal malformations, in malformations by category or in total malformations among all group.
- There were no treatment-related increases in the incidences of individual, visceral or skeletal variations, of variations by category (external, visceral or skeletal) or of total variations.

(See the tables "ORGAN WEIGHTS ", "GESTATIONAL PARAMETERS", "MALFORMATIONS IN FETUSES & LITTERS " and "VARIATIONS IN FETUSES & LITTERS" attached in background material.)

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: See "Details on maternal toxic effects"

Details on maternal toxic effects:
Survival was 100% for all groups during the course of the study.
No gross lesions were seen at necropsy of the study animals
Various clinical signs were observed in the 1000 mg/kg b.w./day group including urogenital wetness, urine stains, urogenital discharge and perinasal encrustation. Urogenital discharge and/or perinasal encrustation were also observed in two dams of the mid dose group.
Gestational body weight and body weight gains and food consumption during the first week of treatment were reduced in the high and mid dose groups.
Absolute and relative liver weight were increased in the high dose group.
No treatment related effects were noted in dams treated with 200 mg / kg b.w./day.
Rats produced signs of maternal toxicity at dose level of 500 and 1000 mg / kg b.w./day.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day
Based on:
act. ingr.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
act. ingr.
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects. Remark: See "Details on embryotoxic / teratogemic effects"

Details on embryotoxic / teratogenic effects:
Fetal morphological observations of the treated groups did not indicate the presence of teratogenic effects at any of the dosage levels tested. (See the table "CESAREAN SECTION OBSERVATIONS" reporteed in "Any other information on results incl. tabels")

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
> 1 000 mg/kg bw/day
Based on:
act. ingr.
Basis for effect level:
other: teratogenicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

The tables reported below have been extracted by the full study report Project number 54-586.

CESAREAN SECTION OBSERVATIONS

Observations

Dose Level [mg PBO/kg/day]

0

200

500

1000

No. Assigned

25

25

25

25

Females gravid

24

21

23

24

Maternal wastage

# died

# sacrificed

# aborted

# early delivery

# non pregnant

 

0

0

0

0

1

 

0

0

0

0

4

 

0

0

0

0

2

 

0

0

0

0

1

corpora lutea/dam

15.3

15.5

15.7

15.0

Total implantants

15.3

14.4

14.5

13.7

Viable implants

14.8

14.1

13.8

13.2

Total resorptions

early

late

 

0.4

0

 

0.2

0

 

0.8

0

 

0.5

0

Dead fetuses

0

0

0

0

Sex ratio (% male fetuses)

50.7

46.5

47.8

50.6

Fetal weight [g]

5.5

5.5

5.4

5.6

Applicant's summary and conclusion

Conclusions:
MATERIALS AND METHODS
In a developmental toxicity study according to Guideline US FIFRA 83-3, groups of 25 mated female Sprague Dawley rats were administered orally by gavage doses of 0, 200, 500, and 1000 mg actual Piperonyl Butoxide /kg bw/day on days 6 through 15 during gestation.
On day 21 of gestation, the fetuses were removed surgically for evaluation.
Dams were observed for clinical signs of toxicity, body weights, food consumption and uterine and liver weights were noted.
Fetuses were dissected from the uterus, counted, sexed, weighed and examined for malformations (visceral and skeletal).

RESULTS AND DISCUSSION
No animals died or were killed in extremis during the study.
Various clinical signs were observed in the 1000 mg/kg b.w./day group including urogenital wetness, urine stains, urogenital discharge and perinasal encrustation. Urogenital discharge and/or perinasal encrustation were also observed in two dams of the mid dose group.
Gestational body weight and body weight gains and food consumption during the first week of treatment were reduced in the high and mid dose groups.
Absolute and relative liver weight were increased in the high dose group.
No treatment related effects were noted in dams treated with 500 or 200 mg Piperonyl Butoxide/ kg b.w./day.
No evidence of fetotoxicity was found, and morphological examination revealed no teratogenic effects at any dose tested.

CONCLUSION
Maternal toxicity was observed in the high dose group. No evidence of fetotoxicity was found, and morphological examination revealed no teratogenic effects at any dose tested. The NOAEL for maternal toxicity was 200 mg actual Piperonyl Butoxide /kg b.w. per day, and that for developmental toxicity was 1000 mg/kg bw per day, the highest dose tested.

LO(A)EL maternal toxic effects = 1000 mg/kg b.w. per day
NO(A)EL maternal toxic effects = 200 mg/kg bw per day
LO(A)EL embryotoxic / teratogenic effects = No effects
NO(A)EL embryotoxic / teratogenic effects = 1000 mg/kg bw per day

Reliability: 1
Deficiencies No