Butyl methacrylate

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OTS 798.5395 (In Vivo Mammalian Cytogenics Tests: Erythrocyte Micronucleus Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Butyl methacrylate

EC Number:

202-615-1

EC Name:

Butyl methacrylate

Cas Number:

97-88-1

Molecular formula:

C8H14O2

IUPAC Name:

butyl methacrylate

Details on test material:

- Supplier: Röhm GmbH
- Name of test material (as cited in study report): n-BUTYL METHACRYLATE
- Physical state: liquid
- Analytical purity: 99,7 % (GC)
- Purity test date: 1998/10/29
- Lot/batch No.: 0078090221M
- Expiration date of the lot/batch: no data

Test animals

Species:

mouse

Strain:

Swiss

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Nossan s.r.l., Corezzana, Italy,
- Age at study initiation: 5-6 weeks
- Weight at study initiation: 26-34 grams for males and 22-27 grams for females
- Assigned to test groups randomly: no data
- Housing: 5/polycarbonate cage, by sexes
- Diet (e.g. ad libitum): Altromin MT diet
- Water (e.g. ad libitum): tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): 55 +/- 10
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

- Vehicle(s)/solvent(s) used: corn oil
- Justification for choice of solvent/vehicle: solubility
- Concentration of test material in vehicle: no data

Duration of treatment / exposure:

Single administration

Post exposure period:

24 and 48 hour sampling time

Doses / concentrations

Remarks:

Doses / Concentrations:
0 (vehicle), 500, 1000 and 2000 mg/kg
Basis:

No. of animals per sex per dose:

Each group consisted of five male and five female animals with the exception of the control and high-dose groups, which included an additional five animals of each sex per group.

Control animals:

yes, concurrent vehicle

Positive control(s):

Mitomycin C
- Route of administration: intraperitoneal
- Doses / concentrations: 2.0 mg/kg bw

Examinations

Tissues and cell types examined:

5 animals/sex/group were sacrificed at the 24-hour sampling time. The  additional animals were sacrificed at the 48-hour sampling time. At least  2000 polychromatic cells per animal were examined for the presence of micronuclei. The ratio of mature to polychromatic erythrocytes was also  determined.  

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION:
A preliminary toxicity study was conducted in which groups of two male and two female mice were dosed once (i.p. injection) with n-butyl  methacrylate at 2000, 1500 and 1000 mg/kg. Clinical signs were observed  in all animals on the day of dosing, but all animals recovered by the  following day. No bone marrow cytotoxicity, as measured by increases in  the NCE/PCE ratio, was observed. The doses for the definitive study were  selected based on this preliminary toxicity study. 

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Five animals per sex from each group were sacrificed at the 24 hour sampling time. The additional animals were sacrificed at the 48 hour sampling time.

DETAILS OF SLIDE PREPARATION:
The femurs were removed and bone marrow cells obtained by flushing with foetal calf serum. The cells were centrifuged and a concentrated suspension prepared to make smears on slides. These slides were air-dried and then stained with May-Gruenwald and Giemsa, and mounted with Eukitt. Three slides were made from each animal.

METHOD OF ANALYSIS:
The slides were randomly coded by a person not involved in the subsequent microscope scoring. The slides were examined under low power (x 16 objective) and one slide from each animal was selected according to staining and quality of smears. At least 2000 polychromatic cells per animal were examined for the presence of micronuclei at high power (x 100 objective, oil immersion). At the same time the numbers of normal and micronucleated normochromatic erythrocytes were also recorded.

Evaluation criteria:

The test substance is considered to induce micronuclei if a statistically significant increase in the micronucleus incidence in polychromatic erythrocytes (at P<0.05) is observed in any treatment group, in the pooled data for both sexes, or for either sex considered separately.

Statistics:

Only counts obtained from polychromatic cells were subjected to statistical analysis. Using the original observations (and not the micronucleus frequencies per 1000 cells), a modified Chi-squared calculation was employed to compare treated and control groups. The degree of heterogeneity within each group was first calculated and where this was significant it was taken into account in the comparison between groups. Variance ratios or Chi-squared values are taken to show the significance of any difference between each treated group and the controls.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Remarks:

but tested up to the limit dose

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

No statistically significant increase in the incidence of micronucleated PCE's over the control value was observed at any dose-level. Slight increases in the ratio of mature to polychromatic erythrocytes, compared to the vehicle control, were seen at the 48 hour sampling time for both male and female animals from the high-dose group.

Any other information on results incl. tables

Following are the results for the male and female animals combined:

                        Incidence of         NCE/PCE
            Dose        micronucleated        Mean
Treatment   Level              PCE              Ratio
           (mg/kg)        Mean      S.E.
-----------------------------------------------------------        
24 hr. Sampling time
-----------------------------------------------------------        
Vehicle (corn oil)
            10           0.5      0.2         1.12
-----------------------------------------------------------
           (ml/kg)
n-butyl methacrylate
           500           0.6      0.2         1.19
n-butyl methacrylate        
          1000           0.9      0.2         0.93
n-butyl methacrylate
          2000           0.6      0.2         1.15
-----------------------------------------------------------
Mitomycin-C  2.0        33.6***   6.5         1.32
-----------------------------------------------------------
48 hr. Sampling time
-----------------------------------------------------------        Vehicle (corn  oil)
           10            0.8      0.3         1.10
-----------------------------------------------------------
          (ml/kg)
n-butyl methacrylate
         2000            1.2      0.3         1.48
-----------------------------------------------------------
*** Incidence significantly greater than control value at p < 0.0001

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
In a valid guideline study, no statistically significant increase in the incidence of micronucleated PCE's over the control value was observed at any dose-level. Slight increases in the ratio of mature to polychromatic erythrocytes, compared to the vehicle control, were seen   at the 48 hour sampling time for both male and female animals from the  high-dose group. The substance was negative for genotoxicity in this test.

Executive summary:

The ability of n-butyl methacrylate to cause chromosomal damagein vivowas investigated in a micronucleus OECD 474 test. Dose-levels for treatment were selected on the basis of a preliminary toxicity test. Male and female Swiss CD-1 mice were dosed once intraperitoneally with vehicle only, corn oil, 2000, 1000 and 500 mg/kg bw n-butyl methacrylate and the positive control Mitomycin-C. Five animals per sex from each group were sacrificed at the 24 hour sampling time. The additional animals were sacrificed at the 48 hour sampling time. Following treatment with n-butyl methacrylate, no statistically significant increase in the incidence of micronucleated PCE's over the control value was observed at any dose-level. Slight increases in the ratio of mature to polychromatic erythrocytes, compared to the vehicle control, were seen at the 48 hour sampling time for both male and female animals from the high-dose group, indicating that the test substance exerted a mild toxic effect on the bone marrow cells. Following treatment with the positive control Mitomycin-C, statistically significant increases in the incidence of micronucleated PCE's over the control values were seen in the positive control group indicating the correct functioning of the test system. It is concluded that n-butyl methacrylate administered intraperitoneally at dose-levels of 2000, 1000 and 500 mg/kg bodyweight to both male and female animals, does not induce micronuclei in the polychromatic erythrocytes of treated mice, under the reported experimental conditions.