Registration Dossier

Diss Factsheets

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:
1989
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study according to OECD guideline 474

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1989
Report date:
1989

Materials and methods

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

Test material

Constituent 1
Chemical structure
Reference substance name:
[(butoxymethylethoxy)methylethoxy]propan-1-ol
EC Number:
259-910-3
EC Name:
[(butoxymethylethoxy)methylethoxy]propan-1-ol
Cas Number:
55934-93-5
Molecular formula:
C13H28O4
IUPAC Name:
[(butoxymethylethoxy)methylethoxy]propan-1-ol
Details on test material:
- Name of test material (as cited in study report): Tripropylene glycole-n-butyl ether
- Molecular formula (if other than submission substance): C13H28O4
- Molecular weight (if other than submission substance): 248

- Physical state: liquid
- Analytical purity:96.12%
- Impurities (identity and concentrations):

- Lot/batch No.: EB880621R004

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, Kingston,NY
- Age at study initiation: 8 week old
- Assigned to test groups randomly: yes
- Housing:singly
- Diet - ad libitum)
- Water - ad libitum)
- Acclimation period:one week


ENVIRONMENTAL CONDITIONS
adequate environmental condition


Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: single oral dose


DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:
Frequency of treatment:
single
Post exposure period:
groups of animals were sacrificed at 3 intervals 24, 48, and 72h after treatment.
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 187.5, 625.0, and 1875.0 mg/kg bw
Basis:

No. of animals per sex per dose:
5 animals per sex per dose per treatment time
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide
- Justification for choice of positive control(s): historical data
- Route of administration: oral
- Doses / concentrations:120mg/kg body weight

Examinations

Tissues and cell types examined:
bone marrow cells for PCE
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: based on the dose range finding test
TREATMENT AND SAMPLING TIMES: at 24, 48, 72h

DETAILS OF SLIDE PREPARATION: At the end of the sacrificed intervals following dosing, the animals were sacrificed by cervical dislocation. Bone morrow samples were obtained from the femurs. Bone marrow was aspirated into fetal bovine serum. After aspiration, the contents of the syringe were transferred into a 1.5 ml centrifuge tube containing 0.5 ml of serum. The cells were resuspended in serum by gentle aspiration using the syringe and needle to centrifuge. After centrifugation supernatant was discarded leaving a small amount of serum covering the pellet. The cell pellet was resuspended using a disposable transfer pipet. Cell smear were prepared on microscope slides using small portions of the cell suspension. The cells were allowed to air dry, fixed in methanol and stained in 5% Giemsa.

METHOD OF ANALYSIS: The slides were coded and scored blindly. One thousand PCE were examined from each animal and the number of micronucleated polychromatic erythrocytes (MN-PCE) was recorded. Micronuclei were identified as darkly stained bodies with sharp contours and varying shapes such as round, almond or ring (Schmid, 1976). The ratio of PCE-NCE in the bone marrow was determined by examining 1000 erythrocytes. The ratio was expressed as PCEx100/PCE+NCE.
Evaluation criteria:
There are several criteria for determining a positive result, such as a dose-related increase in the number of micronucleated cells or a clear increase in the number of micronucleated cells in a single dose group at a single sampling time. Biological relevance of the results should be considered first. Statistical methods may be used as an aid in evaluating the test results.
Statistical significance should not be the only determining factor for a positive response. Equivocal results should be clarified by further testing preferably using a modification of experimental conditions.
A test substance for which the results do not meet the above criteria is considered nonmutagenic
in this test.
Although most experiments will give clearly positive or negative results, in rare cases the data set will preclude making a definite judgement about the activity of the test substance. Results, may remain equivocal or questionable regardless of the number of times the experiment is repeated.
Positive results in the micronucleus test indicate that a substance induces micronuclei which are the result of chromosomal damage or damage to the mitotic apparatus in the erythroblasts of the test species. Negative results indicate that, under the test conditions, the test substance does not produce micronuclei in the immature erythrocytes of the test species.
The likelihood that the test substance or its metabolites reach the general circulation or specifically the target tissue (e.g. systemic toxicity) should be discussed.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative
There were no significant increases in the frequencies of MN-PCE in groups treated with DOWANOL TPnB compared to negative controls. The positive control mice showed significant increases in MN-PCE. Hence, under the experimental conditions used, DOWANOL TPnB was considered negative in the mouse bone marrow micronucleus test.
Executive summary:

DOWANOL TPnB was evaluated in the mouse bone marrow micronucleus test. The micronucleus test is capable of
detecting agents causing chromosomal aberrations and spindle malfunction.  DOWANOL TPnB was mixed with corn oil and administered to CD-1 

(ICR) BR  mice by single oral gavage at dose levels of 0 (negative control), 187.5,  625.0, and 1875.0 mg/kg body weight. 

Groups of animals were sacrificed at three intervals, viz., 24, 48, and 72 hours after treatment. Mice treated with 120mg/kg body weight cyclophosphamide and sacrificed at 24 hour served as positive controls. There were five animals
per sex per dose level per sacrifice time. At the end of the sacrificed intervals following dosing, the animals were sacrificed by cervical dislocation. Bone morrow samples were obtained from the femurs. Bone marrow was aspirated into fetal bovine serum. After aspiration, the contents of the syringe were transferred into a 1.5 ml centrifuge tube containing 0.5 ml of serum. The cells were resuspended in serum by gentle aspiration using the syringe and needle to centrifuge. After centrifugation supernatant was discarded leaving a small amount of serum covering the pellet. The cell pellet was resuspended using a disposable transfer pipet. Cell smear were prepared on microscope slides using small portions of the cell suspension. The cells were allowed to air dry, fixed in methanol and stained in 5% Giemsa.

The slides were coded and scored blindly. One thousand PCE were examined from each animal and the number of micronucleated polychromatic erythrocytes (MN-PCE) was recorded. Micronuclei were identified as darkly stained bodies with sharp contours and varying shapes such as round, almond or ring (Schmid, 1976). The ratio of PCE-NCE in the bone marrow was determined by examining 1000 erythrocytes. The ratio was expressed as PCEx100/PCE+NCE.

Based on study results there were no significant increases in the frequencies of MN-PCE in groups treated with DOWANOL TPnB compared to negative controls. The positive control mice showed significant increases in MN-PCE. Hence, under experimental conditions used, DOWANOL TPnB was considered negative in the mouse bone marrow micronucleus test.