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Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information
None
Link to relevant study records
Reference
Endpoint:
three-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Not specified
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
This data has been reviewed by the World Health Organisation (WHO) have agreed that no effects or dominant lethality was found at either dose level tested. The data is considered accurate and appropriate based on the registrants experience of the substance.
Qualifier:
no guideline followed
Principles of method if other than guideline:
In a three-generation, oral gavage study in rats using CFC-12 (in corn oil) at average doses of 15 and 150 mg/kg per day.
GLP compliance:
not specified
Limit test:
yes
Species:
rat
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
Not specified
Route of administration:
oral: gavage
Vehicle:
not specified
Details on exposure:
In a three-generation, oral gavage study in rats using CFC-12 (in corn oil) at average doses of 15 and 150 mg/kg per day
Details on mating procedure:
Not specified
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
Not specified
Duration of treatment / exposure:
Not specified
Frequency of treatment:
Not specified
Details on study schedule:
Not specified
Remarks:
Doses / Concentrations:
15 or 150 mg/kg
Basis:
no data
No. of animals per sex per dose:
Not specified
Control animals:
not specified
Details on study design:
Male and female rats were given Freon 12 in doses of 15 or 150 mg/kg by intubation. The rats were then bred and evaluated for fertility, corpora lutea, implantation sites, resorption sites, and number of live fetuses per litter.
Positive control:
Not specified
Parental animals: Observations and examinations:
Not specified
Oestrous cyclicity (parental animals):
Not specified
Sperm parameters (parental animals):
Not specified
Litter observations:
Not specified
Postmortem examinations (parental animals):
Not specified
Postmortem examinations (offspring):
Not specified
Statistics:
Not specified
Reproductive indices:
Fertility, corpora lutea, implantation sites, resorption sites, and number of live fetuses per litter.
Offspring viability indices:
Number of live fetuses per litter.
Clinical signs:
not specified
Body weight and weight changes:
not specified
Food consumption and compound intake (if feeding study):
not specified
Organ weight findings including organ / body weight ratios:
not specified
Histopathological findings: non-neoplastic:
not specified
Other effects:
not specified
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
No dominant lethality was found at either dose level.
Dose descriptor:
NOEC
Effect level:
> 150 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Clinical signs:
not specified
Mortality / viability:
not specified
Body weight and weight changes:
not specified
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
not specified
Histopathological findings:
not specified
No dominant lethality was found at either dose level.
Reproductive effects observed:
not specified
Conclusions:
No dominant lethality was found at either dose level. The substance is not considered to be classified for fertility effects.
Executive summary:

In a 3 generation oral study using CFC12 (in corn oil) at average doses of 15 and 150 mg/kg per day, Sherman (1974) found no adverse effects in reproductive capability as measured by the fertility index, gestation index, viability index and lactation index. No classification is applicable.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
150 mg/kg bw/day
Study duration:
chronic
Species:
rat
Quality of whole database:
3-generation study.
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

This data has been reviewed by the World Health Organisation (WHO) have agreed that no effects or dominant lethality was found at either dose level tested.  The data is considered accurate and appropriate based on the registrants experience of the substance.

In a 3 generation oral study using CFC12 (in corn oil) at average doses of 15 and 150 mg/kg per day, Sherman (1974) found no adverse effects in reproductive capability as measured by the fertility index, gestation index, viability index and lactation index. No classification is applicable.

 


Short description of key information:
Fertility effects are discussed.

Justification for selection of Effect on fertility via oral route:
This data is taken from a summary report. Further information is provided below.

Justification for selection of Effect on fertility via inhalation route:
No data

Justification for selection of Effect on fertility via dermal route:
No data

Effects on developmental toxicity

Description of key information
Developmental effects are discussed.
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Not specified
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study report, not conducted to GLP but well documented with methodology and results included.
Qualifier:
no guideline followed
Principles of method if other than guideline:
See "details on exposure" and "details on test animals and environmental conditions" below.
GLP compliance:
not specified
Limit test:
yes
Species:
rat
Strain:
other: Charles River-CD
Details on test animals and environmental conditions:
Seventy-eight pregnant (primigravida) albino rats (Charles River-CD) were received from Charles Rivers Breeding Laboratories in two shipments one day apart. From the time of arrival at Haskell Laboratory through the fifth day of gestation, the animals were housed inn suspended stainless steel wire cages, assigned to three groups, and fed ground Purina Laboratory Chow (GPLC) supplemented with 1% corn oil (CO).
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
On day six of the gestation period, all rats in all groups were given the following diet: 74.25% GPLC + 0.25% CO + 25% Casein + 0.5% vitamin mix. One group, the control, was given 2.0ml of corn oil each day by intragastric intubation for 10 consecutive days; the other two groups were given 2.0ml of Haskell No. 6966 (0.2% solution of Freon 12 in corn oil) or Haskell No. 6967 (2.0% solution of Freon 12 in corn oil), respectively, for the same period of time, i.e., through day 15 of gestation. From day 16 of the gestation period to the time of sacrifice, the control and test rats received their initial basal diet (GPLC + 1% CO).
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
Not specified
Details on mating procedure:
Not specified
Duration of treatment / exposure:
10 days
Frequency of treatment:
Daily
Duration of test:
To day 21 of gestation period.
Remarks:
Doses / Concentrations:
0, 16.6 & 170.9 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
78 rats in total.
Group I (0) - 25
Group II (16.6) - 26
Group III (170.9) - 27
Control animals:
yes, concurrent vehicle
Details on study design:
The animals were sacrificed by chloroform inhalation on the twenty-first day of gestation. Pregnancy was dated by counting as day one the morning on which a copulation plug was found. An incision was made in the abdominal wall and both ovaries and uterine horns were promptly removed, inspected and weighed. The uterus was carefully opened and the foetuses removed. The following observations and measurements were made:
1. Number and location of live foetuses, dead foetuses, and resorption sites in the respective uterine horns.
2. Body weight and crown-rump length of all live foetuses.
3. External examination of all foetuses, from head to tail, under Macro Scope (Ednakite 2½ x) for gross abnormalities.
About two-thirds of the foetuses from each litter were preserved in 95% alcohol for subsequent maceration in 1% aqueous KOH, clearing and staining with Alizarin Red S, and examination to detect skeletal abnormalities. The remaining foetuses were fixed in Bouin’s fluid for free-hand razor-blade sectioning (Wilson Method) and examination for visceral and neural anomalies under the dissecting microscope.
Maternal examinations:
Not specified
Ovaries and uterine content:
An incision was made in the abdominal wall and both ovaries and uterine horns were promptly removed, inspected and weighed. The uterus was carefully opened and the foetuses removed.
Fetal examinations:
Number and location of live foetuses, dead foetuses, and resorption sites in the respective uterine horns. Body weight and crown-rump length of all live foetuses. External examination of all foetuses, from head to tail, under Macro Scope (Ednakite 2½ x) for gross abnormalities.
Statistics:
Not specified
Indices:
Not specified
Historical control data:
The data was also compared with the standard values established for Charles River-CD (ChR-CD) rats.
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
Body Weight: The oral administration of Freon 12 to pregnant rats during the period of organogenesis had no adverse effect on body weight gain
Food Consumption: There were no meaningful differences among the control and test groups of rats with respect to food intake during any of the time periods measured.
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Pregnancy Outcome and Fetal Development: The uteri of all females in all groups were free from gross abnormalities. There was no difference in the numbers of implantation sites, viable foetuses and resorption sites per female between the control group and those given Freon 12. Likewise, there was no significant difference in mean weight and length of foetuses. The smaller weight and length in the control group were due to sacrifice of several females on day earlier (20 days). The data was also compared with the standard values established for Charles River-CD (ChR-CD) rats and were found to be in good agreement.
Malformations and Anomalies: No major gross, skeletal or soft tissue abnormalities were detected. Maternal feeding of Freon 12 did not influence the fetal development. The incidence and type of other minor anomalies were compared with those of the control group and with the standard values for ChR-CD rats. They were found to be similar.
Dose descriptor:
NOAEL
Effect level:
> 170 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: teratogenicity
Dose descriptor:
NOAEL
Effect level:
> 170 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: embryotoxicity
Abnormalities:
not specified
Developmental effects observed:
not specified

TABLE I – AVERAGE BODY WEIGHTS OF PREGNANT RATS RECEIVING FREON 12 DURING ORGANOGENESIS

Group Characteristics

No. of Pregnant Rats

No. of Rats Received

Average Body Weight in Grams on Day of Gestation Period

6

16

21

2 ml corn oil (control)

22/25

199

269

337

2 ml corn oil containing 0.2% Freon 12 (test)

20/26

203

278

345

2 ml corn oil containing 2.0% Freon 12 (test)

25/27

196

271

346

 

TABLE II – AVERAGE FOOD CONSUMPTION DATA OF PREGNANT RATS RECEIVING FREON 12 DURING ORGANOGENESIS

Group Characteristics

No. of Pregnant Rats

No. of Rats Received

Average Daily Food Consumption (g/rat/day)

Control Diet Prior to Day 6

Diet During Dosing Day 6-16

Control Diet Days 16-21

2 ml corn oil (control)

22/25

17.1

17.0

21.0

2 ml corn oil containing 0.2% Freon 12 (test)

20/26

18.7

19.3

22.7

2 ml corn oil containing 2.0% Freon 12 (test)

25/27

17.4

17.8

23.1

 

TABLE III – AVERAGE DOSE OF FREON 12 RECEIVED BY PREGNANT RATS DURING ORGANOGENESIS

Group Characteristics

Average Body Weight In Grams During Day 6-16

Average Dose of Freon 12 (mg/kg/day)

2 ml corn oil (control)

234

-

2 ml corn oil containing 0.2% Freon 12 (test)

241

16.6

2 ml corn oil containing 2.0% Freon 12 (test)

234

170.9

 

TABLE IV – EFFECT OF ORAL ADMINISTRATION OF FREON 12 TO PREGNANT RATS IN THE OUTCOME OF PREGNANCY AND ON VARIOUS PARAMETERS OF FETAL DEVLEOPMENT

 

Groups (mg/kg/day of Freon 12)

Standard Values (Range) for Charles River (CD) Rats (1)

Group I (0)

Group II (16.6)

Group III (170.9)

Number Females Pregnant

22/25 (88.0%)

20/26 (76.9%)

25/27 (92.6%)

90% (62-100)

Number Implantation Sites

201

186

236

 

Number Implantation Sites / Pregnant Female

9.13

9.30

9.44

10.9 (8.6-14.1)

Number Live Fetuses

197

179

231

 

Number Dead Fetuses

0

0

0

 

Number of Live Fetuses/Litter

8.95

8.95

9.24

10.2 (8.2-12.7)

Number Females Showing Complete Resorption

0

0

0

0.3% (0-5.9)

Number Females Showing Premature Birth

0

0

0

0.1% (0-5.9)

Number Females Showing Partial Resorption (Excluding Complete Resorptions)

4 (18.2%)

4 (20.0%)

3 (12.0%)

40.6% (10.5-77.8)

Total Number Resorbed

4 (2.0%)

7 (3.8%)

5 (2.1%)

6.0% (1.2-12.4)

Mean Fetus Weight (g)

3.35

4.19

3.93

3.69 (2.90-4.22)

Mean Fetus Crown-Rump Length (cm)

2.98

3.52

3.36

 

(1) Personal communication; unpublished data obtained from Charles River Breeding Laboratories, Wilmington, Massachusetts for period 1964 -1969.  

 

TABLE V – GROSS EXTERNAL, SKELETAL AND SOFT TISSUE ABNORMALITIES FOUND IN FETUSES FROM MOTHERS ADMINISTERED FREON 12 DURING FETAL DEVELOPMENT

Abnormalities Found

Groups (mg/kg/day of Freon 12)

Standard Values (Range) of Abnormalities for ChR-CD Rats (1)

Group I (0)

Group II (16.6)

Group III (170.9)

1. Gross External

Number Fetuses Examined

197

179

231

32915

a. Small, stunted

1 (0.5%)

0 (0%)

4 (1.73%)

0.3% (0-3.5)

2. Skeletal

Number Fetuses Examined

12

117

154

7127

a. Xiphisternum and one or more sternebrae unoissified

10 (7.7%)

10 (8.5%)

17 (11.0%)

11% (5-30)

b. With 14 rib(s)

15 (11.7%)

9 (7.6%)

12 (7.8%)

21% (6.36)

c. Vertebral bipartite centra

1 (0.8%)

1 (0.9%)

0 (0%)

2% (0-4)

3. Soft Tissue (Visceral and Neural)

Number Fetuses Examined

68

*

77

 

a. Found normal

68

-

77

 

(1) Personal communication; unpublished data obtained from Charles River Breeding Laboratories, Wilmington, Massachusetts for period 1964 -1969.

* Not examined.

Conclusions:
Maternal feeding of CFC-12 was not embryotoxic under the conditions of this experiment.CFC-12 was not teratogenic.
Executive summary:

The present study was undertaken, as part of a comprehensive toxicological program, to evaluate the embryotoxic and teratogenic potential, if any, of CFC-12 when it was administered by intragastric intubation to pregnant rats during the period of fetal organogenesis. 

Gastric intubation of dichlorodifluoromethane (CFC-12) to pregnant rats from day six through day fifteen of their gestation period, at average dose levels of 16.6 and 170.9 mg/kg/day, had no effect on the food intake and the gain in body weight of the mothers.

No clinical signs of toxicity were observed in any of the treated animals. The outcome of pregnancy, measured by the number of implantation sites, resorptions and live foetuses per litter, was not adversely affected by the treatment. The treatment did not affect the embryonal development as measured by weight and crown-rump length of the foetuses. The absence of any major external, skeletal, and soft tissue abnormalities in live foetuses and a similar incidence of spontaneous minor anomalies in the control and treated groups indicate that oral administration for CFC-12 was not teratogenic.

CONCLUSIONS

The absence of any effect on the number of implantation sites, resorptions, live foetuses per litter and mean weight and crown-rump length of the foetuses indicate that maternal feeding of CFC-12 was not embryotoxic under the conditions of this experiment.

Likewise, the absence of any major abnormalities in live foetuses and a similar incidence of spontaneous minor anomalies in the control and treated groups indicate that CFC-12 was not teratogenic.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
170 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

One key study and 3 supporting studies are available for this endpoint.

 

The key study was undertaken, as part of a comprehensive toxicological program, to evaluate the embryotoxic and teratogenic potential, if any, of CFC-12 when it was administered by intragastric intubation to pregnant rats during the period of fetal organogenesis. 

Gastric intubation of dichlorodifluoromethane (CFC-12) to pregnant rats from day six through day fifteen of their gestation period, at average dose levels of 16.6 and 170.9 mg/kg/day, had no effect on the food intake and the gain in body weight of the mothers.

No clinical signs of toxicity were observed in any of the treated animals. The outcome of pregnancy, measured by the number of implantation sites, resorptions and live foetuses per litter, was not adversely affected by the treatment. The treatment did not affect the embryonal development as measured by weight and crown-rump length of the foetuses. The absence of any major external, skeletal, and soft tissue abnormalities in live foetuses and a similar incidence of spontaneous minor anomalies in the control and treated groups indicate that oral administration for CFC-12 was not teratogenic.

 

CONCLUSIONS

The absence of any effect on the number of implantation sites, resorptions, live foetuses per litter and mean weight and crown-rump length of the foetuses indicate that maternal feeding of CFC-12 was not embryotoxic under the conditions of this experiment.

Likewise, the absence of any major abnormalities in live foetuses and a similar incidence of spontaneous minor anomalies in the control and treated groups indicate that CFC-12 was not teratogenic.

 

Supporting studies:

 

In studies of inseminated Wistar albino rats and albino rabbits, Paulet et al. (1974) administered a mixture of 90% CFC-12 (dichlorodifluoromethane) and 10% Freon 11 by inhalation for 2hr/day. Rats were exposed on days 4 to 16 of gestation, and rabbits on days 5 to 20. The mixture was administered in a 20% concentration (200,000 ppm). No indications of any embryotoxic, fetotoxic, or teratogenic changes were found when dams were sacrificed and foetuses removed on day 20 of gestation (rats) or day 30 of gestation (rabbits). No classification is applicable.

 

In a study of a mixture (10% CFC-11 and 90% CFC-12), groups of rats and rabbits were exposed by inhalation on days 4-16 (rats) or days 5-20 (rabbits) of gestation for 2 h/day at a concentration of 989 g/m3(200 000 ppm). No evidence of embryotoxicity, fetotoxicity, or teratogenicity was seen when rats and rabbits were sacrificed at 20 days (rats) or 30 days (rabbits) of gestation. In addition, offspring of the dams that were allowed to deliver naturally showed no evidence of toxicity relative to survival or growth (US EPA, 1983).

 

In a study, groups of 25 to 27 pregnant Charles River rats were given CFC-12 in corn oil by gavage at doses of 16.6 or 179 mg/kg per day on days 6-15 of gestation. Neither dose induced any evidence of embryotoxicity or teratogenicity (Sherman, 1974).

 

In conclusion, CFC-12 did not show any evidence of reproductive or developmental toxicity. No classification is applicable.


Justification for selection of Effect on developmental toxicity: via oral route:
Study report

Justification for selection of Effect on developmental toxicity: via inhalation route:
No data

Justification for selection of Effect on developmental toxicity: via dermal route:
No data

Justification for classification or non-classification

The above studies have all been ranked reliability 2 according to the Klimish et al system. This ranking was deemed appropriate because the studies were not conducted to GLP and are not in compliance with agreed protocols. However these are considered appropriate for use in assessment purposes in accordance with the criteria listed in Annex XI. As such, it is deemed appropriate to apply a weight of evidence approach based on similarity and animal welfare grounds.

The above results triggered no classification under the Dangerous Substance Directive (67/548/EEC) and the CLP Regulation (EC No 1272/2008). No classification for repro effects is therefore required.