Renewable Hydrocarbons C15-C18, branched alkanes

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: According to or similar to guideline study OECD 474.

Data source

Reference

Reference Type:

publication

Title:

Evaluation of the Genetic Toxicity of Middle Distillate Fuels

Author:

R.H. Mckee, M.A. Amoruso, JJ. Freeman, and R.T. Przygoda

Year:

1994

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

GLP compliance:

not specified

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Middle distillate fuels

IUPAC Name:

Middle distillate fuels

Constituent 2

Reference substance name:

kerosines

IUPAC Name:

kerosines

Test material form:

other: low viscosity liquid hydrocarbon

Test animals

Species:

mouse

Strain:

CD-1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
Source: Charles River Breeding Laboratories, Inc.
Sex: Male (65), Female (65)
Age at study initiation: Approximately 9-10 weeks
Weight at study initiation: 23-39g
Housing: Individually
Diet (e.g. ad libitum): Purina Certified Rodent 5002 chow (pellets), ad libitum
Water (e.g. ad libitum): Automatic watering system, ad libitum
Acclimation period: 35d

ENVIRONMENTAL CONDITIONS
Temperature (°F): 68-76
Humidity (%): 40-70%
Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

Corn oil was used. Dose volume did not exceed 10 ml/kg bw.

Details on exposure:

The animals were randomly divided into 5 dose groups: 4 of the dose groups contained 30 mice (15 males and 15 females). The animals in the first 4 groups were treated with corn oil (the vehicle control) or with 1.0, 2.5, or 5.0 g/kg test material. Doses were administered by oral gavage: dosing volumes were 10 mL/kg. Five male and five female mice from each group were sacrificed 24, 48, or 72 hr after treatment, and the bone marrows were isolated and examined for the presence of micronuclei. The last test group contained 10 mice (5 males and 5 females), which were given 40 mg/kg cyclophosphamide by intraperitoneal injection. All of these mice were sacrificed 24 hr after test material administration.

Duration of treatment / exposure:

Animals were sacrificed 24, 48, and 72 hours after dose administration.

Frequency of treatment:

One dose was given of either vehicle control or with 1.0, 2.5, or 5.0 g/kg test material.

Post exposure period:

Animals were sacrificed 24, 48, and 72 hours after dose administration.

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 1.0, 2.5, or 5.0 g/kg
Basis:
actual ingested
oral gavage

No. of animals per sex per dose:

The animals were randomly divided into 5 dose groups: 4 of the dose groups contained 30 mice (15 males and 15 females). The animals in the first 4 groups were treated with corn oil (the vehicle control), or with 1.0, 2.5, or 5.0 g/kg test material. Marrows were isolated and examined for the presence of micronuclei. The last test group contained 10 mice (5 males and 5 females), which were given 40 mg/kg cyclophosphamide.

Control animals:

yes, concurrent vehicle

Positive control(s):

The positive control, cyclophosphamide was administered as a single intraperitoneal injection (40 mg/kg) using water as a carrier.

Examinations

Tissues and cell types examined:

Erythrocytes derived from femur bone marrow.

Details of tissue and slide preparation:

Immediately following the sacrifice of the animals, both femurs were removed and the bone marrow was removed and suspended in fetal bovine serum. After the suspension was centrifuged the pellet was resuspended and smears were prepared (two slides per animal).

Evaluation criteria:

Slides were stained using acridine orange; polychromatic erythrocytes (PCE) stained red/orange, nonchromatic erythrocytes (NCE) are unstained (dull green), and micronuclei stain bright yellow. Slides were evaluated at 400x by fluorescent microscopy. A total of 1000 erythrocytes were counted from each animal, and the numbers of polychromatic (PCE) and normochromatic (NCE) erythrocytes were tabulated. To determine micronucleus (MN) frequency, 1000 PCEs were examined and the number of MN per 1000 PCEs was reported.

Statistics:

Statistical analysis included calculation of means and standard deviations as well as a standard one way analysis of variance (ANOVA) at each time period. When the ANOVA was significant, comparisons of vehicle-treated to dosed group means were made according to Duncans Multiple Range test. A standard regression analysis was performed to test for dose-response relationships. Sexes were analyzed separately.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

None of the five test materials produced an increase in micronucleus frequency regardless of sex or sampling time. Additionally, there was no evidence of bone marrow depression. The positive control (cyclophosphamide) produced a significant increase in micronucleus formation, and the vehicle control values fell within the normal control limits.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
These data indicate that kerosenes are not cytotoxic and are not clastogenic in CD-1 mouse bone marrow cells at doses up to and including 5.0 g/kg of body weight. Classification is not warranted under the new Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP) or under the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.

Executive summary:

Kerosenes were tested in the mammalian bone marrow micronucleus assay using CD-1 mice.  The test materials were tested at 24, 48, and 72 hour intervals following exposure and did not induce a statistically significant decrease in the mean percent of polychromatic erythrocytes or an increase in the mean number of micronucleated polychromatic erythrocytes. Both the positive (cyclophosphamide) and the negative (carrier) controls behaved in an appropriate manner.  These data indicate that kerosenes are not cytotoxic and are not clastogenic in CD-1 mouse bone marrow cells at doses up to and including 5.0 g/kg of body weight.Classification is not warranted under the new Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP) or under the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.