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Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
6 Oct. 1994 - 6 Feb. 1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

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

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
mammalian bone marrow chromosome aberration test

Test material

Constituent 1
Chemical structure
Reference substance name:
Alcohols, C12-14, ethoxylated, sulfates, sodium salts
EC Number:
500-234-8
EC Name:
Alcohols, C12-14, ethoxylated, sulfates, sodium salts
Cas Number:
68891-38-3
Molecular formula:
not applicable, UVCB
IUPAC Name:
Alcohols, C12-14, ethoxylated, sulfates, sodium salts

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 Italia S.p.A., Calco, Como, Italy
- Age at study initiation: 5 - 6 weeks
- Weight at study initiation: 25 - 32 g (males); 20 - 27 g (females)
- Assigned to test groups randomly: yes, immediately after arrival at test facility
- Fasting period before study: yes, overnight
- Housing: 5 animals/cage, separated by sexes, in clear polycarbonate cages (type 2b: Techniplast) with a stainless steel mesh lid and floor; each cage equiped with absorbent bedding which was inspected daily and changed as necessary.
- Diet: Altromin MT, ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATE: 26 Oct. 1994

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
Vehicle(s)/solvent(s) used: injectable grade sterile distilled water
- Lot/batch no.: 22143-lB (Don Baxter S.p.A., Trieste, italy)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Fresh solutions of the test substance were prepared for each day's work; solutions were prepared on a weight/volume basis. Solutions of the test substance, following corrections of active constituent were prepared in injectable grade sterile distilled water.
Duration of treatment / exposure:
not applicable
Frequency of treatment:
single treatment
Post exposure period:
Test groups: 10, 24 and 48 h
Positive control group: approx. 26 h
Doses / concentrationsopen allclose all
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Remarks:
corrected for a.i. content of test material
Dose / conc.:
2 000 mg/kg bw/day (actual dose received)
Remarks:
corrected for a.i. content of test material; represents the recommended limit dose for administration periods < 14 days according to OECD TG 475
No. of animals per sex per dose:
Test groups: 5 for each sampling time (10, 24 and 48 h)
Positive control group: 5
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide (50 mg/kg bw)

Examinations

Tissues and cell types examined:
Tissue: femoral bone marrow
Cell type: femoral bone marrow cells
Details of tissue and slide preparation:
SAMPLING TIMES
Test groups: 10, 24 and 48 h after treatment
Positive control group: 26 h after treatment

CRITERIA FOR DOSE SELECTION:
Range finding study performed to find the maximum tolerated dose.

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Following treatment and prior to sample collection, animals are injected intraperitoneally with 4 mg/mL colchicine and samples are collected 3 h thereafter. Cells are harvested from the bone marrow, swollen, fixed and stained, and analysed for chromosomal aberrations.

DETAILS OF SLIDE PREPARATION:
Slides were fixed with a methanol:acetic acid solution, air-dried and stained with Giemsa.

METHOD OF ANALYSIS:
50 metaphase spreads per animal were examined microscopically at high magnification for the presence of chromosomal aberrations. Only metaphases containing 40 chromosomes are scored for aberrations. The number of chromosomes, the specific types and numbers of aberrations are recorded. The Vernier readings of aberrant or equivocal metaphases are recorded.

OTHER:
The mitotic index is calculated. This is based on the number of metaphases observed per 1000 cells and is expressed as a percentage.
Evaluation criteria:
A test substance is considered to have clastogenic properties if a statistically significant increase in the incidence of cells bearing aberrations is observed at any test point, excluding gaps. The lesions observed must be consistent with those expected at the sampling time at which the increases are observed. The internal consistency of the findings within groups are also taken into account.
Statistics:
The experimental unit investigated in the present study is the proportion of cells from an animal which exhibit chromosomal aberrations. It is therefore assumed that the data has a binomial error structure, each cell out of a total of 'n' for each animal either bearing or not-bearing aberrations.
A logistic-linear model is fitted to the data adding the factors 'Sex', 'Treatment' and 'Sex-Treatment Interaction' in turn. The change in deviance with each factor is compared with the Chi-squared distribution to determine the level of significance. The deviance associated with the residual term is compared with the Chi-squared distribution to determine the goodness-of-fit of the logistic-linear model. This allows the construction of a table similar to a classical analysis of variance.
Data from in vivo cytogenetic assays often contain extra-binomial variation, which may result in part from animal-to-animal variation, or from the responses of different populations of cells within animals. This situation will be highlighted by a significant lack of fit of the logistic-linear model. When this is the case, an allowance for the extra-binomial variation can be made using a generalised 'heterogeneity factor' to weight the logistic-linear model variables.
The statistical analysis is performed using the values for the total number of aberration-bearing cells observed for each animal, excluding gaps from consideration.

Results and discussion

Test results
Key result
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: 250, 500, 1000 and 2000 mg/kg bw
- Clinical signs of toxicity in test animals: No clinical signs were observed at any dose level tested.
- Evidence of cytotoxicity in tissue analysed: No reduction in the mitotic index was observed in any treatment group.

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative

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