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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
The study was conducted in accordance with OECD Test guideline No 422 and with the principles of GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
No further details.
Specific details on test material used for the study:
- Purity: 99.96%

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Crj:CD(SD)IGS rats used but no details provided regarding supplier
- Age at study initiation: (P) x wks; (F1) x wks No data
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g No data
- Fasting period before study: No data
- Housing: No data

- Diet (e.g. ad libitum):
- Water (e.g. ad libitum):
- Acclimation period:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data
- Humidity (%): No data
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data

IN-LIFE DATES: From: not stated To: not stated

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5% sodium carboxymethylcellulose solution
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Justification for use and choice of vehicle (if other than water):0.5% sodium carboxymethylcellulose solution used as suspending agent
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
No information provided
Details on mating procedure:
- M/F ratio per cage: 1:1.
Duration of treatment / exposure:
Males were dosed for 42 days prior to mating; the females were treated for 14 days prior to mating and then until day 4 of lactation. The female satellite animals were treated or 42 days. The recovery period for males and satellite females was 14 days. The males were terminated on day 43 of treatment (day 15 of recovery) and the females were terminated on day 5 of lactation, satellite females on day 15 of recovery and the offspring were terminated 4 days after birth.
Frequency of treatment:
Once daily for up to 42 days
Duration of test:
6 weeks
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
8 mg/kg bw/day (actual dose received)
Dose / conc.:
40 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
twelve; an additional five rats per sex were allocated to the satellite groups for retention through the recovery phase.
Control animals:
yes, concurrent vehicle
Details on study design:
No further details provided.
As a screening study this investigation was limited to the parental generation and F1 offspring only. The repeated adminstration phase for males (42 days) and females (14 days) prior to mating provides some subacute toxicity data and the treatment of females through gestation gives some reproductive and developmental toxicity information.

The males and females were dosed for up to 42 days by oral gavage and this subacute exposure was used to address the short-term toxicology endpoints. Clinical signs, bodyweight gain, food consumption haematology, clinical chemistry and urinalysis parameters were recorded together with a limited functional observation battery, organ weights and macroscopic and microscopic examinations at termination.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: at weekly intervals - assessment in homecage, then in observers hands and then outside the homecage
Assessments included posture, sleeping, locomotion,vocalisations, tremors and convulsions, response to capture and handling, salivation, grading of heart beat, body temperature, exophthalmus and pupil size, exhibition of any discoloration of fur,skin or lacrimation. Outside the cage the assessment of posture, grooming, vocalisations, occurrence of straub tail, gait, tremor, convulsion pilo-erection and palpebral opening; exploration and respiratory rate and any exhibiion of stereotypy or bizarre behaviour.

A functional assessment was completed on the final day of treatment to assess various reflex responses.

BODY WEIGHT: recorded on days 1, 7, 14, 21, 28, 35 and 42 of treatment and days 1, 7, and 14 of recovery for males.
Females were weighed on days 1, 7, and 14 of treatment; days 0, 7, 14 and 20 of pregnancy and days 0 and 4 of lactation
Bodyweights and bodyweight gains are reported.

Food consumption was recorded for the males and for the satellite females (treated at 200 mg/kg) on days 1-2, 7-8, 14-15, 29-30, 35-36 and 41-42 and then days 6-7 and 13-14 of the recovery phase.

Urinalysis was performed on Day 31 and 32 for males and females respectively. Parameters assessed included colour, turbidity, pH, protein, glucose, ketone and bilirubin levels, occult blood, urobilinogen and presence in urinary sediment of crystals by incidence and type/shape and urinary specific gravity.

Clinical pathology in the form of standard haematology and biochemical parameters were assessed on Day 43 of treatment and on Day 15 of the recovery phase for male rats and on Day 5 of lactation for the females.

Organ weights were recorded on Day 43 of treatment and on Day 15 of the recovery phase for male rats and on Day 5 of lactation for the females.


Necropsy was completed on termination of treatment or after completion of the recovery phase or on day 5 of lactation. Macroscpic findings were recorded at each scheduled necropsy. Histopathological findings were documented for tissues sampled following each of the scheduled necropsy events. The list of tissues examined was in accordance with the test guideline.

The reproductive performance of the parents was recorded for the twelve pairings per treatment group; the number of pairs copulating provided the copulation index and the number of pregnant females was used to calculate the fertility index. the mean number of days from pairing to copulation was also calculated

All females examined for oestrous cyclicity . The number of rats showing a 4 day cycle in the pre-treatment phase was recorded, together with number with 5-day cycle. Mean cycle duration was calculated
Ovaries and uterine content:
No data
Fetal examinations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring on day 0 of lactation:
number of pups, stillbirths, live births, birth index and live birth index

Bodyweights recorded on day 0 and 4 of lactation

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities

Necropsy of live pups surviving to lactation day 4- examination of external and visceral changes
Statistics:
No data
Indices:
Number of newborn, delivery index, number of live newborn, birth index and live bith index calculated on day 0 of lactation. On day 4 of lactation the number of live pups, viability index and sex ratio were calculated.
Historical control data:
No data

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
Oral administration of 4,4'-biphenyldiol at dose levels of 8, 40 or 200 mg/kg bw/day did not cause death or a moribund condition in any animals. None of the scores obtained during detailed clinical observations differed in a biologically significant manner between the control and the compound-treated groups.
No apparent changes were observed in general clinical conditions, except that urine in the 200 mg/kg-treated group became cloudy with elapse of time after excretion. Urinalysis was performed on treatment days 31 and 32 in male and female rats, respectively, calcium oxalate-like urinary crystal sediments were found in males given 200 mg/kg and in females given 40 or 200 mg/kg. Turbidity was enhanced at dose levels of 40 or 200 mg/kg, and urinary specific gravity was decreased in these females. These changes in urine parameters were not found when urinalysis was performed 11 days after cessation of the treatment (on day 11 of recovery) in either sex for the satellite animals.
Body weight, weight gain and food consumption were not affected by treatment at any dose level in either sex. No animals showed any abnormality in functional parameters after the final treatment.
At necropsy on termination of the treatment, no apparent effects of the compound were found on haematological or blood-biochemical examination at any dose level of the compound in either sex.

The reproductive and developmental toxicity indices evaluated in this screening study indicated no adverse effects on the dams. No effects were apparent in the female oestrous cycle either during the pre-treatment, mating or treament phases. The reproductive indices included the numbers of pairings examined, copulating and the number of resulting pregnant females and the length of pairing time required before copulation. There were no changes between control and treated groups.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
External malformations:
no effects observed
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There were no adverse effects of administration on offspring viability, morphology or growth.
Pup development indices were evaluated to assess the duration of gestation, numbers of corpora lutea and implantations, the numbers of pups born, numbers of live born and sex ratio for the liveborn pups and survival and viability of pups to day 4 of lactation was also evaluated . None of the comparisons between treated and control groups revelaed any significant changes in these indices or any statistically significant effects. Similarly no effects were apparent when newborn or neonate (lactation day 4) pup weights were assessed.

Morphological evaluations for external and visceral changes showed no effects on the live pups at birth or those that survived to lactation Day 4. The dead pups examined (3 controls and 1, 6 or 12 pups in the 8, 40 or 200 mg/kg bw/day groups) also had no external and visceral changes .

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
>= 200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No effect observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Any other information on results incl. tables

Combined repeat dose and reproductive/developmental toxicity screening test for 4,4’-biphenyldiol

 

4,4’-biphenyldiol mg/kg bw/day

 

0

8

40

200

No. of females examined

12

12

12

12

No. of females showing 4-day cycle in pre-treatment phase

12

12

12

12

No. of females showing 4-day cycle in treatment phase

12

11

12

12

No. of females showing 4 and 5-day cycle in treatment phase

0

1

0

0

Mean length of oestrous cycle in days

4.0±0.0

4.0±0.1

4.0±0.0

4.0±0.0

Number of vaginal oestrous during mating period

1.0±0.0

1.0±0.0

1.0±0.0

1.0±0.0

 

 

4,4’-biphenyldiol mg/kg bw/day

 

0

8

40

200

No. of pairs examined

12

12

12

12

No. of pairs copulated

11

11

12

12

Copulation index

91.7

91.7

100.0

100.0

No. of pregnant females

11

11

10

12

Fertility index

100.0

100.0

83.3

100.0

Number of pairing days
 to copulation

2.5±1.1

2.6±1.2

2.7±0.8

2.7±1.2

 

  Development of pups up to Day 4 of lactation

 

4,4’-biphenyldiol mg/kg bw/day

 

0

8

40

200

No. of pregnant females

11

11

10

12

No. of pregnant females with live neonates

11

11

10

12

Gestation index

100

100

100

100

Gestation length in days

22.4±0.5

22.4±0.5

22.0±0.0

22.4±0.5

Number of corpora lutea

15.9±1.6

16.5±1.1

16.2±1.2

15.2±1.5

Number of implantations

15.5±1.7

14.3±3.6

16.0±1.6

14.7±1.7

Implantation Index

97.1±4.3

86.9±21.6

98.7±4.2

96.7±5.3

 

 

 

 

 

Day 0 of lactation

 

 

 

 

Number of newborn

14.5±2.3

13.8±4.0

15.1±1.7

14.0±1.8

Delivery index

93.2±6.9

94.6±12.2

94.4±4.8

95.5±5.2

Number of live newborn

14.3±2.3

13.8±4.0

14.6±1.6

12.8±2.9

Birth index

92.0±7.6

94.6±12.2

91.5±8.1

88.8±20.1

Live birth index

98.7±3.0

100.0±0.0

97.0±7.5

93.3±20.8

Sex ratio on day 0

54.8±14.5

49.4±17.4

52.5±15.1

51.6±14.7

 

 

 

 

 

Day 4 of lactation

 

 

 

 

Number of live pups

13.9±2.1

13.6±4.0

14.5±1.5

12.4±4.2

Viability index

97.7±4.0

98.8±2.6

99.4±2.0

91.7±28.9

 

 

 

 

 

Sex ratio on day 4

54.4±14.9

49.3±17.9

52.1±15.9

50.8±15.2


The vehicle control was 5 ml/kg of 0.5% CMC Na solution.
Gestation index
      (number of pregnant females with live newborns/number of pregnant females) x 100,%
Implantation index
 (number of implantations/number of corpora lutea) x 100,%
Delivery index
        (number of newborns/number of implantations) x 100,%
Birth index
             (number of live newborns/number of implantations) x 100,%
Live birth index
      (number of live newborns/number of newborns) x 100,%
sex ratio (day 0)
     (number of male live newborns / number of live newborns) x 100,%
viability index
        (number of live pups on day 4 of lactation/ number of live newborns) x 100,%
sex ratio (day 4)
     (number of male live pups on day 4 of lactation / number of live pups on day 4 of lactation) x 100,%

Bodyweight of pups up to Day 4 of lactation

 

4,4’-biphenyldiol mg/kg bw/day

 

0

8

40

200

Day 0 of lactation

 

 

 

 

Number of live newborn

 

 

 

 

male

7.7±2.1

6.9±3.0

7.6±2.1

6.5±2.2

female

6.5±2.5

6.9±3.3

7.0±2.6

6.3±2.7

 

 

 

 

 

Birthweight of live newborn

 

 

 

 

Males

6.8±0.8

6.9±0.8

6.5±0.5

6.6±.0.8

females

6.6±0.6

6.6±0.7

6.0±0.4

6.2±0.8

 

 

 

 

 

Day 4 of lactation

 

 

 

 

Number of live pups

 

 

 

 

Males

7.5±2.3

6.8±3.1

7.5±2.2

6.8±2.0

females

6.4±2.2

6.8±3.2

7.0±2.6

6.7±2.5

Birthweight of pups

 

 

 

 

Males

10.6±1.7

10.7±1.1

10.2±1.1

10.8±1.0

females

10.3±1.4

10.4±1.4

9.8±1.0

10.0±0.9

Morphological findings of pups

 

4,4’-biphenyldiol mg/kg bw/day

 

0

8

40

200

Dead pups

 

 

 

 

Number of dead pups examined

3

1

6

12

External observations

3

1

6

12

Visceral observations

0

0

5

1

No. of pups with external changes

0

0

0

0

No. of pups with visceral changes

0

0

0

0

 

 

 

 

 

 

 

 

 

 

Live pups at birth (Day 0)

 

 

 

 

Number of live neonates examined

157

152

146

154

Number with external changes

0

0

0

0

At necropsy on Day 4

 

 

 

 

Number of pups examined

153

150

145

149

No. of pups with external changes

0

0

0

0

No. of pups with visceral changes

0

0

0

0



Applicant's summary and conclusion

Conclusions:
Treatment with up to 200 mg/kg bw/d did not affect early offspring development. The no observed adverse effect level (NOAEL) for repeat dose toxicity of 4,4'-biphenyldiol is considered to be 40 mg/kg bw/d. In the absence of any effects on the parameters measured. the NOAEL for developmental toxicity is considered to be 200 mg/kg/ bw/d.
Executive summary:

A combined repeat dose and reproductive/developmental toxicity screening test was conducted in rats according to the OECD Test Guideline 422 with 4,4'-biphenyldiol. No effects on developmental parameters were observed at any dose level. In the absence of any effects on developmental parameters, a NOAEL for developmental toxicity of 200 mg/kg bw/d can be determined. A NOAEL for general toxicity of 40 mg/kg bw/d can be determined based on liver effects at the highest dose level. Effects of treatment on urinalysis parameters seen at 40 and 200 mg/kg bw/d are not considered to be of toxicological significance.