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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
August 20 – September 24, 1985
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study performed according to US EPA guidelines.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EPA OPP 84-2
Deviations:
no
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Naphthalene; 5601-56-1
- Substance type: Polycyclic aromatic hydrocarbon
- Physical state: White flake
- Analytical purity: Not provided
- Impurities (identity and concentrations): Not provided
- Lot/batch No.: #J-272
- Expiration date of the lot/batch: Not provided
- Stability under test conditions: Stipulated stable
- Storage condition of test material: Room temperature

Test animals

Species:
mouse
Strain:
CD-1
Sex:
male/female
Details on test animals and environmental conditions:
CD-1. Charles River Breeding Laboratory, Wilmington, MA

SOURCE:
- Age at study initiation: 8 ½ week
- Weight at study initiation: Males 28 – 35 g; females 26 – 29 g
- Fasting period before study: Not fasted
- Housing: 5 per cage per sex per dose group
- Diet (e.g. ad libitum): Wayne Rodent Blox, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS – Definitive study
- Temperature (°C): 21 -22°C
- Humidity (%): 64 – 65%
- Air changes (per hr): not provided
- Photoperiod (hrs dark / hrs light): 12 hr dark /12 hr light

Administration / exposure

Route of administration:
intraperitoneal
Vehicle:
- Vehicle(s)/solvent(s) used: vegetable oil, Mazola corn oil
- Concentration of test material in vehicle: Range: 250,166.6, 150, 50, 25 mg/ml; Definitive 25 mg/ml
- Amount of vehicle (if gavage or dermal): 10 ml/kg
- Type and concentration of dispersant aid (if powder): None

Duration of treatment / exposure:
Single injection
Frequency of treatment:
Once
Post exposure period:
30, 48, 72 hours
No. of animals per sex per dose:
5 per sex per dose
Control animals:
yes
Positive control(s):
- Control substance: triethylenemelamine, Lot # 25238, Polyscience, Inc., in 0.9% saline
- Route of administration: intraperitoneal
- Doses / concentrations: 0.5 mg/kg

Examinations

Tissues and cell types examined:
Bone marrow cells from femur
Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION: Range-finder
- TREATMENT AND SAMPLING TIMES (in addition to information in specific fields): 250 mg/kg sampled at 30, 48 and 72 hours
- DETAILS OF SLIDE PREPARATION: Cell suspension dropped on slide and stained with Giemsa
- METHOD OF ANALYSIS: Microscopic. 1000 PCE counted for presence of micronucleated PCE. Data expressed as number of micronucleated PCE versus total normal PCE per 1000 total PCE per animal.
Evaluation criteria:
Micronuclei are uniform and darkly staining typically round bodies in the cytoplasm of PCE. Inclusions in PCEs which are reflective, improperly shaped or stained, or which are not in the focal plane of the cell are judged to be artefacts and are not scored as micronuclei. Cells containing more than one micronucleus are only scored once.
Statistics:
One-tailed “t” test to evaluate pairwise treatment groups with negative control. Statistical significance was judged at p< 0.05 and p< 0.01 levels.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
not determined
Toxicity:
yes
Remarks:
Signs of toxicity at 250 mg/kg.
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: 250 – 5000 mg/kg
- Solubility: 250 – 500mg/kg (25, 50 mg/ml)
- Clinical signs of toxicity in test animals: Decrease in body tone, abnormal gait, death at 500 mg/kg.
- Evidence of cytotoxicity in tissue analyzed: Not determined
- Rationale for exposure: 250 mg/kg determined to be the MTD
- Harvest times: 30, 48 and 72 hours
- High dose with and without activation: 250 mg/kg; activation not relevant

RESULTS OF DEFINITIVE STUDY
- Types of structural aberrations for significant dose levels (for Cytogenetic or SCE assay):
- Induction of micronuclei (for Micronucleus assay): None
- Ratio of PCE/NCE (for Micronucleus assay): Depression of PCE/NCE ratio at 72 hour sacrifice.
- Appropriateness of dose levels and route: Maximum tolerated dose (MTD)
- Statistical evaluation: “t” test negative

Any other information on results incl. tables

Animal Number

Control

Naphthalene (5601-56-1)

 

Corn Oil

TEM

250 mg/kg

 

10 ml/kg

0.5 mg/kg

30 hours

48 hours

72 hours

Male

1

0

60

1

2

2

2

1

63

0

0

1

3

3

65

1

1

1

4

1

55

1

0

1

5

2

72

0

0

1

Female

1

0

55

1

1

0

2

2

56

0

0

0

3

0

38

1

1

1

4

2

75

0

2

1

5

2

73

1

0

0

Mean

1.30 ­± 1.06

61.2 ­± 11.09

0.60 ­± 0.52

0.70 ­± 0.82

0.80 ± 0.63

“t“ value

 --

16.997**

1.878*a

1.414

1.281

*, ** Denotes statistical significance at p<0.05 and 0.01 respectively

a Lower than control mean

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Naphthalene is not a clastogen.
Executive summary:

In a preliminary dose-range finding study, naphthalene was administered intraperitoneally to 5 groups (2 males and 2 females per group) of CD-1 mice at dose levels of 250, 500, 1000, 3000, and 5000 mg/kg of body weight. Animals were observed for pharmacotoxic signs for up to 72 hours after dose administration. Pharmacotoxic signs were observed at all levels evaluated. All animals died at the 4 highest levels while animals at 250 mg/kg all survived treatment. Due to the severity of the signs and the mortality observed in the study, 250 mg/kg was selected as the dose for the Micronucleus Test as an estimate of the maximum tolerated dose. In the Micronucleus Test, three groups of ten animals (5 males and 5 females/group) were given single doses by intraperitoneal injection at 250 mg/kg and sacrificed at 30, 48, and 72 hours. Similar groups, dosed with the positive control, triethlyenemelamine (TEM), and negative control, corn oil, were evaluated concurrently. Slides were prepared from the bone marrow of the femurs and stained with Geimsa. Coded slides were scored for the number of polychromatic erythrocytes (PCE) with micronuclei in 1000 PCE per animal. The ratio of polychromatic to normochromatic erythrocytes per 1000 erythrocytes per animal was determined. The results for naphthalene were negative in the Micronucleus Test at a dose level of 250 mg/kg at all of the time intervals evaluated. These findings are based upon the inability of the test substance to produce a statistically significant increase in the number of micronuclei in 1000 polychromatic erythrocytes per animal in the treated groups versus the negative control group.