Octafluorocyclobutane

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July 2012 - March 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5395 (In Vivo Mammalian Cytogenetics Tests: Erythrocyte Micronucleus Assay)

GLP compliance:

yes

Type of assay:

other: peripheral blood samples in mammalian erythrocytes micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Perfluoroethane

EC Number:

200-939-8

EC Name:

Perfluoroethane

Cas Number:

76-16-4

Molecular formula:

C2F6

IUPAC Name:

hexafluoroethane

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Name of test material : perfluoroethane (CAS 76-16-4) named H-30188 in the study
- Lot/batch number of test material: 1011MT0161 ; 1110CW0033 and 0911MT0144
- Purity: 99.99% according to Certificate of Analysis (COA - appendix A of the study)

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under storage conditions: Stable based on analyses conducted by the sponsor. The test substance appeared to be stable under the storage conditions.
- Stability under test conditions: The test substance appeared to be stable under the conditions of the study; no evidence of instability was observed.
- Solubility and stability of the test substance in the dispersant/vehicle/test medium: Test substance is a gas miscible with the houseline air used in the study.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: No treatment prior to testing
- Preliminary purification step: No

Test animals

Species:

rat

Strain:

other: Crl:CD(SD)

Remarks:

nulliparous, non-pregnant rats

Details on species / strain selection:

The Crl:CD(SD) strain was chosen because extensive background data are available from the literature, the supplier, and previous studies at
DuPont Haskell. This strain is also considered suitable relative to longevity, hardiness, and incidence of spontaneous disease.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories International, Inc., Kingston, New York
- Age at Arrival: Approximately 51 days
- Age at Start of Inhalation: Approximately 70 days
- Weight on day after arrival: 198-240 grams (males); 148-183 grams (females)
- Weight at start of inhalation: 317-445 grams (males) and 201-269 grams (females)
- Assigned to test groups randomly: Yes, under following basis: Computerized, stratified randomization so that no statistically significant group mean body weight differences occur within a sex.
- Housing: Individually in solid bottom caging with bedding, and enrichment as appropriate; sexes on separate racks except during cohabitation (for reproduction group)
- Cohabitation: Mating pairs (females placed in males’ cages) on a 1:1 basis in solid bottom caging with bedding and enrichment. Fourteen days after the first day of cohabitation, females without evidence of copulation were housed singly.
- Diet: PMI® Nutrition International, LLC Certified Rodent LabDiet® 5002 ad libitum except when fasted.
- Water: Tap water ad libitum
- Acclimation period: approximately 20 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-26ºC (68-79ºF)
- Humidity (%): Relative humidity of 30-70%
- Photoperiod (hrs dark / hrs light): Artificial (fluorescent light) on an approximate 12-hour light/dark cycle (except during eye examinations and motor activity).

Administration / exposure

Route of administration:

inhalation: gas

Vehicle:

- Vehicle used: Chamber atmospheres were generated by dilution of the tested substance in houseline air and oxygen.
- Justification for choice of vehicle: The test material is a gas. The mode of exposition is via inhalation.

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- Exposure mode :
During exposure, animals were individually placed in stainless steel, wire-mesh modules and exposed whole-body inside the exposure chamber, except during the cohabitation period (for reproduction group), when animals were housed as mating pairs and exposed in the same manner.

- Atmosphere Generation:
H-30188 (test material) vapor and supplemental oxygen were metered into a 1-liter 3-neck mixing flask by Brooks model 5850 E or 5850 I mass flow controllers. The mixture left the mixing flask and entered the glass transfer tube where chamber air supply was added to the mixture by Brooks model 5850 E or 5851 I mass flow controllers. The gas mixture then entered the top of the exposure chamber through a turret. The air-control atmosphere was similarly generated in a different room without the H-30188 vapor.
Test atmospheres were exhausted from the bottom of each chamber through MSA filters using vacuum pumps and discharged into the fume hood.

- Chamber Distribution of Test Substance:
Uniform distribution of the test substance in the 150-L and 350-L exposure chambers was determined.

Duration of treatment / exposure:

approximately 20 days of exposure

Frequency of treatment:

6 hours/day, 5 days/week

Post exposure period:

Not specified

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 ppm

Doses / concentrations 2

Dose / conc.:

2 500 ppm

Doses / concentrations 3

Dose / conc.:

10 000 ppm

Doses / concentrations 4

Dose / conc.:

50 000 ppm

No. of animals per sex per dose:

5 rats/sex/dose

Control animals:

yes

Positive control(s):

No

Examinations

Tissues and cell types examined:

Approximately 6-8 drops of blood (peripheral blood samples) was collected by tail vein venipuncture from the 5 rats/sex/group. There were two blood collections, one following the fourth exposure (test day 3) and following the last exposure before scheduled euthanasia.
Cell types examined were immature erythrocytes and normochromatic erythrocytes.

Details of tissue and slide preparation:

No information

Evaluation criteria:

Results considered positive if:
- statistically significantly increase mean MN-RETs (micronucleated reticulocytes) observed at one or more concentrations of test substance compared to concurrent negative control values
- statistically significant dose-response increase in MN-RETs observed

Statistics:

Yes

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

not specified

Negative controls validity:

valid

Any other information on results incl. tables

Micronucleus Evaluation for Male Rats – Subchronic Subset

	Group 1 0 ppm	Group 2 2,500 ppm	Group 3 10,000 ppm	Group 4 50,000 ppm
RET(%)
3-Day	1.63 0.34 (5)	a	a	1.32 0.25 (5)
28-Day	1.34 0.17(5)	a	a	1.14 0.32 (5)
MN-RET (%)
3-Day	0.10 0.06 (5)	a	a	0.08 0.02 (5)
28-Day	0.11 0.03 (5)	a	a	0.09 0.04 (5)

Data arranged as:

Mean

Standard deviation (Number of values included in calculation)

a = Group not evaluated at this time point.

There were no statistically significant differences at p<0.05

 

Micronucleus Evaluation for Female Rats – Subchronic Subset

	Group 1 0 ppm	Group 2 2,500 ppm	Group 3 10,000 ppm	Group 4 50,000 ppm
RET(%)
3-Day	1.15 0.13 (5)	1.08 0.22(5)	1.10 0.41(5)	1.07 0.18 (5)
28-Day	1.35 0.26(5)	a	a	1.36 0.31 (5)
MN-RET (%)
3-Day	0.06 0.01 (5)	0.09 0.04 (5)	0.10 0.03 (5)	0.09 0.05 (5)
28-Day	0.08 0.02 (5)	a	a	0.07 0.02 (5)

Data arranged as:

Mean

Standard deviation (Number of values included in calculation)

a = Group not evaluated at this time point.

There were no statistically significant differences at p<0.05

 

Applicant's summary and conclusion

Conclusions:

No statistically significant increases in the frequency of MN-RETs compared to the negative control group were observed in any evaluated test substance treated group of male or female rats at any time point. There were no statistically significant test substance-related decreases in %RET. The negative control group exhibited a response consistent with the %MN-RETs historical control data.

Executive summary:

The objective of this study was to evaluate the potential subchronic, reproductive, and genetic effects from repeated-inhalation exposure to the test substance, H-30188, when administered to male and female rats during premating, cohabitation, gestation, and lactation up through day 3.

No statistically significant increases in the frequency of MN-RETs compared to the negative control group were observed in any evaluated test substance treated group of male or female rats at any time point. There were no statistically significant test substance-related decreases in %RET. The negative control group exhibited a response consistent with the %MN-RETs historical control data.