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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:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1994-12-21 to 1995-03-01
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study is classified as reliable without restrictions because it was GLP compliant and was conducted according to OECD 474 guidelines.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
This substance is very similar with regard to health endpoints to the substance being registered.
- Name of test material (as cited in study report): Alkane 4
- Substance type: Poly alpha olefin (1-dodecene trimer, hydrogenated)
- Physical state: Liquid
- Analytical purity: Not reported
- Lot/batch No.: C1527-04-4
- Stability under test conditions: Stable at 24 hours
- Storage condition of test material: Room temperature
- Other: Clear colourless liquid

Test animals

Species:
mouse
Strain:
CD-1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (U.K.) Ltd
- Age at study initiation: 5 to 7 weeks
- Weight at study initiation: Male - 21 to 27 grams; Female - 20 to 24 grams
- Assigned to test groups randomly: Yes, under following basis: animals were selected at random and given a number unique within the study by ear punching and the number written on a colour coded cage card.
- Fasting period before study: No
- Housing: Groups of 5 by sex in steel mesh bottom polypropylene cages
- Diet (e.g. ad libitum): Rat and Mouse Expanded Diet No. 1 (Special Diets Services Limited, Essex, U.K.) ad libitum
- Water (e.g. ad libitum): Ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 22°C
- Humidity (%): 47 to 54%
- Air changes (per hr): 15 per hour
- Photoperiod (hrs dark / hrs light): 12 hours dark/12 hours light

Administration / exposure

Route of administration:
intraperitoneal
Vehicle:
- Vehicle(s)/solvent(s) used: Arachis oil
- Justification for choice of solvent/vehicle: Not reported
- Concentration of test material in vehicle: 125, 250, and 500 mg/mL depending on the concentration tested
- Lot/batch no. (if required): 014316
- Purity: Not reported
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Test material was freshly prepared as required as a solution at the appropriate concentration in arachis oil (The volume administered to each animal was calculated according to its bodyweight at the time of dosing).
Duration of treatment / exposure:
72 hours but 1 animal from each dose group was sacrificed at 24 hours, 48, hrs, and 72 hours post-dosing
Frequency of treatment:
Single intraperitoneal dose
Post exposure period:
72 hours
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
1250 mg/kg
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
2500 mg/kg
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
5000 mg/kg
Basis:
analytical conc.
No. of animals per sex per dose:
5/sex/dose
Control animals:
yes
Positive control(s):
- Positive control: Cyclophosphamide
- Justification for choice of positive control(s): Not reported
- Route of administration: Oral
- Doses / concentrations: 50 mg/kg

Examinations

Tissues and cell types examined:
Only bone marrow smears prepared from extracted femurs were analyzed in the study.
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: Based on the lack of cytotoxicity observed in the highest dose tested in the range-finding study

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): One group of mice from each dose group was killed by cervical dislocation 24 hours following treatment, a second a t 48 hours and a third a t 72 hours. The vehicle control groups were killed 24, 48 and 72 hours following dosing and positive control group animals were killed 24 hours following dosing.

DETAILS OF SLIDE PREPARATION: Immediately following sacrifice one femur was dissected from each animal aspirated with foetal calf serum and bone marrow smears prepared following centrifugation for 30 seconds at approximately 6000 rpm and re-suspension. The smears were air dried, fixed in absolute methanol and stained in May-Grunwald/Giemsa, dried and cover slipped using mounting medium.

METHOD OF ANALYSIS: Stained bone marrow smears were coded and examined ‘blind’ using light microscopy at x1000 magnification. The incidence of micronucleated cells per 1000 polychromatic erythrocytes (PCE- blue stained immature cells) per animal was scored. In addition, the number of
normochromatic erythrocytes (NCE - pink stained mature cells) associated with 1000 polychromatic erythrocytes was counted; these cells were also scored for incidence of micronuclei. The ratio of polychromatic to normochromatic erythrocytes was calculated together with appropriate group mean values for males and females separately and combined.
Evaluation criteria:
A positive mutagenic response is demonstrated when a statistically significant dose-related increase in the number of micronucleated polychromatic erythrocytes is observed for either the 24, 48, or 72-hour kill times when compared to their corresponding control group.

A positive response for bone marrow toxicity is demonstrated when the dose group mean polychromatic to normochromatic ratio is shown to be statistically significantly lower than that of the concurrent vehicle control group.
Statistics:
Data was analyzed following a √(x + 1) transformation using Student's t-test (two tailed) and any significant results were confirmed using the one way analysis of variance.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: 1250, 2500, and 5000 mg/kg
- Solubility: Not reported
- Clinical signs of toxicity in test animals: None observed
- Evidence of cytotoxicity in tissue analyzed: No
- Rationale for exposure: No premature deaths or clinical observations were observed in animals dosed with the test material up to the maximum recommended-dose level of 5000 mg/kg. Therefore the maximum recommended dose (5000 mg/kg) along with 2500 and 1250 mg/kg were selected as the dose levels for use in the main study.

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): There was a small statistically significant increase in the frequency of micronucleated PCEs in the 24-hour 5000 mg/kg test material dose group when compared to the concurrent vehicle control group. The response seen was not part of a dose relationship and was within the historical range for 24-hour vehicle control groups. Therefore, the response was considered to be spurious and of no toxicological significance.
- Ratio of PCE/NCE (for Micronucleus assay): There was no statistically significant change in the PCE/NCE ratio in any of the test material dose groups when compared to their concurrent vehicle control groups.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Alkane 4 was considered to be non-genotoxic under the study conditions
Executive summary:

Justification for Read Across

Several criteria justify the use of the read across approach to fill data gaps for the poly alpha olefin 1-decene trimer and tetramer hydrogenated (CAS number 68649-12-7) using alkane 4 (1-dodecene, trimer, hydrogenated) as an analog. Both of these compounds are poly alpha olefins, i.e., highly branched isoparaffinic chemicals produced by oligomerization of 1-octene, 1-decene, and/or 1-dodecene. They have similarities in chemical structure and physiochemical properties. The literature also indicates that alkanes with 30 or more carbon atoms are unlikely absorbed when administered orally. In turn, studies indicate that they have similar health effect endpoints. Both of these poly alpha olefins have low order acute toxicity and repeated dose toxicity. Additionally gene mutations assays in bacterial cells, as well as in vitro chromosomal aberrations in mammalian cells assays demonstrate no evidence of genotoxicity regardless of metabolic activation. In vivo chromosomal aberrations assays in rats and mice also show no mutagenic potential regardless of metabolic activation. There do not appear to be any toxicological differences between 1-decene trimer and tetramer hydrogenated and alkane 4. Therefore, read across between 1-decene trimer and tetramer hydrogenated and alkane 4 can be justified.

In a CD-1 mouse bone marrow micronucleus assay, 5 mice/sex/dose were treated via intraperitoneal injection with Alkane 4 at doses of 0, 1250, 2500, or 5000 mg/kg bw. Bone marrow cells were harvested at 24, 48, and 72 hours post-treatment. The vehicle was Arachis Oil.

 

Although there was a statistically significant increase in micronucleated cells at 24 hours at 5000 mg/kg; an increase was not observed at 48 or 72 hours; the results were within the historical control response and there was no dose response. The positive control induced the appropriate response. There was no significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow after any treatment time.

 

This study received a Klimisch score of 1 and is classified as reliable without restrictions because it was GLP compliant and was conducted according to OECD 474 guidelines.