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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:
15 Aug - 11 Nov 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
(adopted 22 January 2001)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
other: Japenese Ministry of Agriculture, Forestry and Fisheries Testing guidelines for Toxicology studies, 12 NohSan No 8147, (24 November, 2000)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
The Department of Health of the Government of the United Kingdom
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Lanolin Alcohol
- Physical state: yellow solid
- Analytical purity: >90%
- Batch No.: 0000664004
- Expiration date of the batch: 19 Feb 2014
- Storage condition of test material: room temperature in the dark
- Other: Label: SUPER HARTOLAN-PA-(RB) 0000664004 BATCH

Test animals

Species:
rat
Strain:
other: Sprague-Dawley Crl:CD (SD) IGS BR
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Limited, Margate, Kent, UK
- Weight at study initiation: 214 to 299 g
- Housing: The animals were housed individually in solid-floor polypropylene cages with stainless steel lids furnished with softwood flakes (Datesand Ltd., Cheshire, UK)
- Diet: pelleted diet (Rodent 2018C Teklad Global Certified Diet, Harlan UK, Oxon, UK); ad libitum
- Water: drinking water; ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±3
- Humidity (%): 50±20
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 18 Aug 2013 To: 05 Sept 2013

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: Arachis oil BP
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was prepared at the appropriate concentrations as a solution in Arachis oil BP. The stability and homogeneity of the test item formulations were previously determined by Harlan Laboratories Ltd., Shardlow, UK Analytical Services (Harlan Laboratories Ltd., Project Number 41301173). Results showed the formulations to be stable for at least twenty days. Formulations were therefore prepared once and stored at approximately +4°C in the dark.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples were taken of each test item formulation and were analyzed for concentration of Lanolin Alcohols at Harlan Analytical Laboratory, Shardlow. The test item concentration in the test samples was determined by HPLC with UV detection using an external standard technique. The results indicate that the prepared formulations were within ± 2% of the nominal concentration.
Details on mating procedure:
- Impregnation procedure: purchased timed pregnant
- The day that positive evidence of mating was observed was designated Day 0 of gestation.
Duration of treatment / exposure:
Day 5 - 19 of gestation
Frequency of treatment:
daily
Duration of test:
15 days
No. of animals per sex per dose:
24 (except for the intermediate treatment group: Only 23 animals were used in the intermediate treatment group as upon delivery of the animals, one female was found to have a damaged eye. This female was unsuitable to be used on the study and was humanely killed.)
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were chosen based on previous toxicity data.
- Other: The oral route was selected as the most appropriate route of exposure, based on the physical properties of the test item, and the results of the study are delieved to be of value in predicting the likely toxicity of the test item to man.

Examinations

Maternal examinations:
CLINICAL OBSERVATIONS: Yes
- Time schedule: Following arrival, all animals were examined for overt signs of toxicity, ill-health or behavioral changes once daily during the gestation period. Additionally, during the dosing period, observations were recorded immediately before and soon after dosing and one hour post dosing.

BODY WEIGHT: Yes
- Time schedule for examinations: on Day 3 of gestation (before the start of treatment) and on Days 5, 6, 7, 8, 11, 14 and 17 of gestation. Body weights were also recorded for animals at terminal kill (Day 20).

FOOD CONSUMPTION: Yes
- Time schedule for examinations: for each individual animal at Day 3, 5, 8, 11, 14, 17 and 20 of gestation.

WATER CONSUMPTION: Yes
- Time schedule for examinations: daily by visual inspection of the water bottles for any overt changes

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation Day 20
- All animals were subjected to a full external and internal examination and any macroscopic abnormalities were recorded.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number, position and type of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Placental weight: Yes
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter
- Skeletal examinations: Yes: half per litter
- Visceral examinations: Yes: half per litter
Statistics:
Female body weight change, food consumption and gravid uterus weight: Bartlett’s test for homogeneity of variance and one way analysis of variance, followed by Dunnett’s multiple comparison test or, if unequal variances were observed, on alternative multiple comparison test.
All caesarean necropsy parameters and fetal parameters: Kruskal-Wallis non-parametric analysis of variance; and a subsequent pairwise analysis of control values against treated values using the Mann-Whitney ‘U’ test, where significance was seen.
Fetal evaluation parameters, including skeletal or visceral findings: Kruskal-Wallis nonparametric analysis of variance and Mann-Whitney ‘U’ test.
Probability values (p) are presented as follows:
p<0.001 ***
p<0.01 **
p<0.05 *
p≥0.05 (not significant)
Indices:
Pre- and post-implantation losses
Historical control data:
Historical control data from several former embryofetal development toxicity studies were provided, giving information on skeletal findings, visceral examination, and reproductive data.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: Observed effects on body weight, food consumption, clinical signs, pre-implantation loss and number of corpora lutea. However, these effects were considered not to represent an adverse effect or to be incidental and unrelated to treatment.

Details on maternal toxic effects:
Mortality: There were no unscheduled deaths (table 3).

Clinical Observations: Females treated with 1000 and 300 mg/kg bw/day showed isolated incidences of increased salivation between Days 6 and 13 (table 2). No such effects were detected in females treated with 100 mg/kg bw/day. Observations of this nature are commonly observed following the oral administration of an unpalatable test item formulation and in isolation are considered not to be of toxicological significance. One female treated with 1000 mg/kg bw/day also showed fur loss between days 16 and 20 (table 2). Observations of this nature are commonly observed in isolation are not considered to be related to test item toxicity. Therefore, no treatment-related effects were found.

Body Weight: No adverse effects on body weight development were detected. Statistical analysis of the data did not reveal any significant intergroup differences (table 3).

Food Consumption: Females treated with 1000 mg/kg bw/day showed a statistically significant increase in food consumption between Days 14 and 17 (table 4). The increase in food consumption is not considered to represent an adverse effect of treatment. Furthermore, intergroup differences on body weight were considered of no toxicological significance.

Water Consumption: Daily visual inspection of water bottles did not reveal any overt intergroup differences.

Pathological examinations: No macroscopic abnormalities were detected.

Females treated with 1000 and 300 mg/kg bw/day showed a statistically significant reduction in the number of corpora lutea (table 5). This intergroup difference was not dose-related and was considered to be incidental and unrelated to treatment due to ovulation and mating occurring prior to the administration of the test item. Females treated with 1000 mg/kg bw/day also showed a statistically significant reduction in pre-implantation loss (table 5). A reduction in this parameter is considered not to represent an adverse effect of treatment therefore the intergroup difference was considered of no toxicological importance.

Effect levels (maternal animals)

open allclose all
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Litter Data and Litter Placenta and Fetal Weights:
There were no treatment-related effects on in utero offspring survival, as assessed by the mean numbers of early or late resorptions, live litter size and post-implantation losses. There was also no adverse effect in sex ratio.
For all dose groups, there were no significant treatment-related trends in the proportion of fetuses (or litters) with evidence of external, visceral or skeletal anomalies (tables 6, 7 and 8). The type of external, visceral and skeletal anomalies, were those commonly observed for this type of study. There were no findings that were considered to represent any known malformations. Statistical analysis of the data did not reveal any significant differences.
Therefore, no treatment-related effects were detected in the uterine parameters examined, in fetal viability or in growth and development.

Effect levels (fetuses)

Key result
Remarks on result:
not determinable because of methodological limitations

Fetal abnormalities

Key result
Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Table 1: Summary of female performance

 

Control group

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Initial group size

24

24

23

24

Pregnant animals

22

24

20

24

 

Table 2: Summary incidence of daily clinical observations

Dose level [mg/kg bw/day]

Number of animals

Clinical observations

Number of animals showing effect (day of observation)

0

24

no abnormalities detected

-

100

24

no abnormalities detected

-

300

23

Increased salivation

3 (6, 9)

1000

24

Increased salivation

15 (6-13)

Generalized fur loss

1 (16-20)

 

Table 3: Group mean values on pregnant females

Parameter

Control group

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Number of dams examined

22

24

20

24

Mortality of dams [%]

0.0

0.0

0.0

0.0

Body weight gain [g]

Day 5–20 of gestation

121.3±15.5

125.6±15.6

124.4±16.4

126.8±14.7

Gravid uterus weight [g]

82.3±9.7#

80.3±14.6

80.3±9.3

81.2±11.2

#: n=21 for gravid uterus weight. Gravid uterus weight not recorded for one female in error.

 

 

Table 4: Group mean food consumption values

Dose level [mg/kg bw/day]

Number of dams examined

Food consumption (g/rat/day) between days of gestation

3-5

5-8

8-11

11-14

14-17

17-20

0

22

25.6±2.0

21.0±2.2

23.0±2.6

24.7±1.9

25.7±2.2

25.2±2.1

100

24

25.0±2.2

21.8±3.4

23.3±2.6

25.2±3.4

26.1±2.6

27.2±5.0

300

20

25.5±2.5

21.5±2.2

23.0±3.3

25.2±2.8

26.9±3.6

26.5±3.9

1000

24

25.4±3.1

22.0±2.0

24.7±2.3

26.2±2.0

27.9±2.6**

27.4±3.0

** Significantly different from control: p** < 0.01

 

Table 5: Group mean litter data values

Parameter

Control group

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Number of corpora lutea

14.9±1.5

14.4±1.8

13.0±1.5***

13.5±2.0**

Number of implants

13.5±1.7

12.8±2.6

12.5±1.2

13.1±1.8

Total number of live implants

12.8±2.4

12.6±2.7

12.2±1.4

12.5±2.0

Total number of embryonic/fetal deaths

1.0±1.6

0.3±0.5

0.3±0.4

0.6±1.6

Pre-implantation loss [%]

8.8±7.9

12.0±13.7

4.0±4.9

2.8±5.5**

Post-implantation loss [%]

5.8±12.0

1.0±2.7

2.1±3.7

4.0±10.4

Total number of litters

22

24

20

24

Mean fetal weight[g]

4.073±0.156

4.157±0.196

4.217±0.156

4.166±0.262

Mean placental weight[g]

0.559±0.049

0.553±0.067

0.577±0.037

0.582±0.056

** Significantly different from control: p** < 0.01

*** Significantly different from control: p*** < 0.001

Table 6: Summary incidence of fetal external findings

Parameter

0 (Control)

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Number of foetuses examined

281

303

244

301

Small fetus

0

1

1

1

Omphalocele

1

0

0

0

Pale

0

0

1

1

Total number of foetuses with fetal external findings (% of foetuses with fetal external findings)

1 (0.4)

1 (0.4)

2 (0.8)

2 (0.7)

 

Table 7: Summary incidence of fetal visceral findings

Parameter

0 (Control)

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Number of foetuses examined

146

157

128

157

Total number of foetuses with fetal visceral findings (% of foetuses with fetal external findings)

51 (33.5)

53 (34.3)

34 (26.5)

70 (45.1)

 

Table 8: Summary incidence of fetal skeletal findings

Parameter

0 (Control)

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Number of foetuses examined

135

146

117

144

Total number of foetuses with fetal skeletal findings (% of foetuses with fetal external findings)

117 (88.6)

123 (84.7)

96 (81.5)

114 (78.8)

 

NOTE: a fetus may appear in more than one category

 

Applicant's summary and conclusion