Oligomerisation products of 2,2'-[(1-methylethylidene)bis(4,1-phenyleneoxymethylene)]bisoxirane with acrylic acid and lauric acid

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

Study period:

April 03 2007 to august 07, 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study following OECD guideline 474 without any deviations

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

BADGEDA, Ebecryl ® 3700

IUPAC Name:

BADGEDA, Ebecryl ® 3700

Details on test material:

- Name of test material (as cited in study report): BADGEDA, Ebecryl ® 3700, CT# 879-07
- Source: Cytec Surface Specialties (Smyrna, USA)
- Physical state: Transparent, light-brown, very viscous liquid
- Analytical purity: ca. 63%
- Lot/batch No.: N1140127
- Date received: March 21, 2007
- Expiration date of the lot/batch: March 19, 2008
- Storage condition of test material: Room temperature
- Stability under test conditions: Stable for 24 hours under room temperature

Test animals

Species:

mouse

Strain:

other: CD-1® (ICR) BR

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan, Frederick, USA
- Age at study initiation: 8 weeks
- Weight at study initiation: Preliminary study: 32.9-39.4 g (males) or 25.8-29.4 g (females); Main study: 32.1-40.0 g (males)
- Assigned to test groups randomly: Yes, animals randomized using a computer program
- Housing: Individually housed in sanitary polycarbonate cages
- Diet (e.g. ad libitum): PMI Certified Rodent Diet® #5002, ad libitum
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 64-79 °F
- Humidity (%): 30-70%
- Air changes (per hour): 10/hour
- Photoperiod (hours dark / hours light): 12 hours dark/12 hours light

IN-LIFE DATES: From: April 12, 2007 To: April 19, 2007

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: Corn oil
- Concentration of test material in vehicle: 25 or 50 mg/mL
- Amount of vehicle (if gavage): 20 mL/kg
- Lot/batch no. (if required): 12-455
- Source: Welch, Holme, & Clarke
- Storage: Refrigerated at 0-10 °C

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Test material was emulsified with corn oil to give stock solution which was then diluted further to give the desired dose levels; formulations were held in a waterbath at ~60 to 70 °C prior to dosing and stirred during the dosing procedure to prevent emulsion breakdown

Duration of treatment / exposure:

24 or 48 hours

Frequency of treatment:

Once for dose levels of 500 and 1000 mg/kg bw, and twice, approximately one hour apart, for the dose level of 2000 mg/kg bw (1000 mg/kg/dose, BID)

Post exposure period:

Not applicable

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

500 other: mg/kg bw/day (actual ingested)

Doses / concentrations 2

Dose / conc.:

1 000 other: mg/kg bw/day (actual ingested)

Doses / concentrations 3

Dose / conc.:

2 000 other: mg/kg bw/day (actual ingested)

No. of animals per sex per dose:

- Range-finding study: 3 mice/sex/dose
- Main study: Five male mice/dose

Control animals:

yes, concurrent vehicle

Positive control(s):

Cyclophosphamide
- Lot no.: 076K1050
- Storage: Refrigerated at 0-10 °C
- Route of administration: Oral (gavage)
- Doses / concentrations: 80 mg/kg bw

Examinations

Tissues and cell types examined:

- Hind limb bones (tibias) were removed for marrow extraction and the prepared slides were examined for polychromatic erythrocytes (PCEs), normochromatic erythrocytes (NCEs) and total erythrocytes

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: Dose selection was based on a preliminary range-finding test conducted on 3 mice/sex/dose at 500, 1000 and 2000 mg/kg bw. No mortality and/or toxic signs were recorded after 2 days of exposure.

TREATMENT AND SAMPLING TIMES: Hind limb bones (tibias) were removed for marrow extraction from five surviving animals in each treatment and control group at 24 or 48 hours after exposure.

DETAILS OF SLIDE PREPARATION: Bone marrow cells extracted, preparations spread on slides and air-dried. The slides were fixed in methanol, stained with May-Grunwald solution and Giemsa and coded.

METHOD OF ANALYSIS: Slides were scanned to determine the frequency of micronuclei in 2000 polychromatic erythrocytes (PCEs) per animal. In addition, PCE:NCE ratio was determined in a population of 500 erythrocytes/animal.

Evaluation criteria:

- Criteria for a positive response: Detection of a statistically significant increase in micronucleated PCEs for at least one dose level, and a statistically significant dose-related response.
- Test article that does not induce both of these responses is considered negative.
- Biological relevance of the results should be considered first.

Statistics:

- Statistical analysis was performed using ANOVA/Program Trend computer system.
- ANOVA followed by Dunnett’s t-test performed on untransformed proportions of cells with micronuclei per animal and on untransformed PCE:NCE ratios when the variances were homogeneous. Ranked proportions were used for heterogeneous variances.

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose range: 500-2000 mg/kg bw
- Solubility: Solubility limit in corn oil: 50 mg/mL
- Clinical signs of toxicity in test animals: No
- Evidence of cytotoxicity in tissue analyzed: Not examined
- Rationale for exposure: No appropriate toxicity data were available

RESULTS OF DEFINITIVE STUDY
- Frequency of micronuclei in polychromatic erythrocytes and PCE:NCE ratio: See table 1

Any other information on results incl. tables

Table 1: Micronucleus Assay - Summary Table Treatment Dose Harvest Time (hours) % Micronucleated PCEs Mean of 2000 per Animal ± S.E.(Males) Ratio PCE:NCE Mean ± S.E.(Males) Controls Vehicle Corn Oil 20 mL/kg 24 0.04 ± 0.02 0.51 ± 0.06 48 0.03 ± 0.02 0.35 ± 0.01 Positive CP 80 mg/kg 24 2.77 ±0.28* 0.55 ± 0.1 Test Article 500 mg/kg 24 0.04 ± 0.03 0.43 ± 0.05 1000 mg/kg 24 0.06 ± 0.02 0.32 ±0.07** 2000 mg/kg^a 24 0.02 ± 0.01 0.59 ± 0.06 48 0.05 ± 0.02 0.4 ± 0.08 * Significantly greater than the corresponding vehicle control, p ≤ 0.01. ** Significantly less than the corresponding vehicle control, p ≤ 0.05. CP = Cyclophosphamide; PCE = Polychromatic erythrocyte; NCE = Normochromatic erythrocyte ^aDue to solubility limit, the high dose was dosed at 1000 mg/kg BID (~1 hr apart) to achieve a dose of 2000 mg/kg.

Applicant's summary and conclusion

Conclusions:

Under the test conditions, the test substance was not genotoxic in the mouse bone marrow micronucleus test.

Executive summary:

In a mouse bone marrow micronucleus test, performed according to OECD guideline 474, in compliance with GLP, CD-1® (ICR) BR male mice (5/dose) were given a single oral (gavage) dose of the test substance Bisphenol A diglycidyl ether diacrylate (BADGEDA) in corn oil at concentrations of 500 and 1000 mg/kg bw and twice, approximately one hour apart, at a concentration of 2000 mg/kg bw (1000 mg/kg/dose, BID). Bone marrow was extracted after 24 or 48 h of exposure and the slides prepared were scanned to determine the frequency of micronuclei in 2000 polychromatic erythrocytes (PCEs) per animal. In addition, the number of PCEs and normochromatic erythrocytes (NCEs) in a population of 500 erythrocytes was determined as a measure of cytotoxicity. A preliminary range-finding test was also conducted on 3 mice/sex/dose at 500, 1000 and 2000 mg/kg bw and animals were observed for 2 days. In this previous test, no mortality and/or toxic signs were recorded. No statistically significant increases in the frequency of micronucleated PCEs were observed at any dose level. A statistically significant decrease in the PCE:NCE ratios at a dose level of 1000 mg/kg bw was observed but was not considered biologically significant as the highest dose of 2000 mg/kg bw did not show a similar trend. Under the test conditions, the test substance was not genotoxic in the mouse bone marrow micronucleus test (Yong, 2007).