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Toxicity to reproduction: other studies

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Administrative data

Endpoint:
toxicity to reproduction: other studies
Type of information:
experimental study
Adequacy of study:
key study
Study period:
24 September to 10 December 1984
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: A well-reported GLP study, conducted according to good scientific principles; considered acceptable for assessment

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985
Reference Type:
secondary source
Title:
Unnamed
Year:
2008

Materials and methods

Test guideline
Qualifier:
no guideline required
Principles of method if other than guideline:
The study was conducted to investigate the findings of a previous study (IRDC, 1985) which had shown a high proportion of pup mortality 2-3 weeks after birth (probably as a result of internal haemorrhaging) from mothers fed 6250 ppm Cereclor S52, and to establish the pattern of exposure (in utero and/or lactational) required to cause the effects seen.
GLP compliance:
yes
Type of method:
in vivo

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Cereclor S52
- Substance type: technical product
- Physical state: pale yellow liquid
- Analytical purity: 100%
- Impurities (identity and concentrations): no stabiliser present
- Composition of test material, percentage of components: a C14-17 chlorinated paraffin (52% chlorinated)
- Purity test date: no data
- Lot/batch No.: Y01063/003/003
- Expiration date of the lot/batch: no data
- Stability under test conditions: no data
- Storage condition of test material: diet containing test material stored at -20oC

Test animals

Species:
rat
Strain:
other: Charles River COBS (Wi) [Wistar]
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS- Source: Charles River UK Ltd, Margate, Kent, UK- Age at study initiation: (P) 10 wks- Weight at study initiation: (P) Males: 295-322 g; Females: 202-230 g; (F1) Both sexes 5.92-6.4 g- Fasting period before study: no data- Housing: stainless steel cages- Diet (e.g. ad libitum): conventional, ad libitum- Water (e.g. ad libitum): filtered tap water, ad libitum- Acclimation period: 10 daysENVIRONMENTAL CONDITIONS- Temperature (°C): 20 ± 2- Humidity (%): minimum 47- Air changes (per hr): no data- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
DIET PREPARATION- Rate of preparation of diet (frequency): prepared in 3 batches on 17 September and 1 and 30 October 1984- Mixing appropriate amounts with (Type of food): mixing with a small amount of diet then adding this to the bulk of the powdered (meal) diet- Storage temperature of food: -20oC
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
measured by extraction and titration
Duration of treatment / exposure:
from 4 weeks pre-mating until either days 10-17 of pregnancy or throughout pregnancy and lactation (see "Any other information on materials and methods incl. tables")
Frequency of treatment:
daily in feed
Duration of test:
from 4 weeks pre-mating until either days 10-17 of pregnancy or throughout pregnancy and lactation (see "Any other information on materials and methods incl. tables")
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:6250 ppmBasis:nominal in diet
Remarks:
Doses / Concentrations:6020, 4618 and 6110 ppm for batches 1, 2 and 3, respectivelyBasis:analytical conc.
Remarks:
Doses / Concentrations:3125 mg/kg bw/dayBasis:actual ingested
No. of animals per sex per dose:
50 males and 100 females during pre-mating period135-150 pups per group with approximately equal numbers of each sex
Control animals:
yes, concurrent no treatment
Details on study design:
See "Any other information on materials and methods incl. tables" for details of the treatment regime.Blood was taken from one pup per litter from all groups on days 3, 4, 5, 8 and 11 and from two pups per litter at termination (day 22) and assayed for the clotting factors VIII and X. Where the volume was sufficient, blood was also analysed for prothombin and platelets measured.
Statistics:
The treated group mean was compared to the control group mean using a two-sided Student's t-test for the following parameters: body weight gain, food consumption and food utilisation during the pre-mating period, male and female fertility indices, length of gestation, pre-coital interval, female body weight gain during pregnancy, pup (litter) body weight, live births, sex distribution, litter size, percentage litter deaths, offspring litter weights, percentage factor VIII, percentage factor X and platelet count

Results and discussion

Effect levels

Dose descriptor:
LOAEL
Effect level:
ca. 3 125 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Feeding a C14-17 chlorinated paraffin (52% chlorination) at about 3125 mg/kg bw/day (only tested dose) to mothers during the lactational period resulted in increased mortality in the pups likely due to internal haemorrhaging.

Observed effects

No treatment-related adverse effects were seen in the pre-mating or pregnancy stages of the study and there was no significant differences in litter size between groups.The offspring from groups in which the lactating mothers were given the test substance (Groups 2 and 4) showed higher pup mortality than groups 1,3 and 5 were the mothers were untreate during lactaion). This was first seen between lactational days 6 and 11, and associated with internal haemorrhages. Between lactational days 6-11 three times more mortalities were seen in treated pups reared by treated mothers (group 4) than control pups reared by treated mothers (group 2). Between lactational days 11-22 more deaths occurred in both groups with slightly more in group 2 (77%) compared to group 4 (67%). A reduction in factor X of 45 and 63% occurred in group 2 and 4, respectively, and increased prothrombin times were detected in the blood of these two groups only.Body weight gain in group 4 showed a statistically significant decrease by day 22.

Applicant's summary and conclusion

Conclusions:
Cereclor S52 (a C14-C17 chlorinated paraffin; 52% chlorinated) caused an increased mortality in the neonatal offspring of rats when given in the diet of lactating dams at about 3125 mg/kg bw/day. No increased mortality was evident when Cereclor S52 exposure occurred only during the pre-mating period and pregnancy. No effects on fertility or foetal viability were detected.
Executive summary:

A study was conducted to investigate whether the findings of a previous study, in which a high proportion of pups whose mothers had been fed Cereclor S52 died 2-3 weeks after birth, was due to in utero and/or lactational pup exposure.

In a good quality study, to GLP, Cereclor S52 (a C14-17 chlorinated paraffin; 52% chlorinated) was given to groups of male and female rats in the diet at 0 or 6250 ppm (about 0 or 3125 mg/kg bw/day) during the pre-mating period and to the females throught pregnancy, or up to days 10-17 of pregnancy or throughout pregnancy and lactation. Groups of control animals received no test substance. After confirmation of mating, the pregnant females were allocated to five groups, as follows:

1. Females fed control diet rearing their own pups.

2. Females fed Cereclor S52 rearing pups from group 3 control females.

3. Females fed control diet rearing pups fostered from group 2 treated females.

4. Females fed Cereclor S52 rearing their own pups.

5. Females fed Cereclor S52 up to days 10-17 of pregnancy, rearing their own pups whilst fed control diet.

Blood samples were obtained from one pup/litter on days 3, 4, 5, 8, and 11, and two pups/litter on day 22 (the day of sacrifice) post-partum and analysed for clotting factors VIII and X. Prothrombin times were also measured at these timepoints and platelet counts on days 11 and 22. Samples of breast milk were taken from lactating dams of groups 1, 2 and 4 on day 14 post‑partum only, and analysed for MCCP.

No treatment-related adverse effects were detected during the pre-mating or pregnancy phases of the study. Between 11 and 13 litters were delivered for each group.

The offspring from groups 2 and 4, in which the lactating mothers were given Cereclor S52, showed higher pup mortality, first seen between post-natal days 6 and 11, and associated with internal haemorrhages in the pups. This suggests that lactational exposure was responsible for the mortalities. A decrease in clotting factor X was seen only in pups exposed lactationally; the decrease was 45 and 63% in group 2 and 4, respectively.

Based on the decrease observed in factor X, the investigators proposed that Cereclor S52 was either transferred to the breast milk resulting in a disruption of the vitamin K-dependent clotting system in the pups and/or that the test substance caused less vitamin K to be available in the breast milk due to treatment-related effects in the mothers.

In conclusion, feeding Cereclor S52 at about 3125 mg/kg bw/day to mothers during the lactational period resulted in increased mortality in the pups likely due to internal haemorrhaging. No such an effect was reported when Cereclor S52 was administered prior to, and during pregnancy, in the absence of lactational exposure.