1-Butanamine, N-butyl-, reaction products with polyethylene-polypropylene glycol ether with trimethylolpropane (3:1) acrylate

Genetic toxicity: in vivo

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:

2012/2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

adopted 1997

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted May 2008

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5395 (In Vivo Mammalian Cytogenetics Tests: Erythrocyte Micronucleus Assay)

Version / remarks:

adopted August 1998

GLP compliance:

yes (incl. QA statement)

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

propylidynetrimethanol, ethoxylated, esters with acrylic acid and its reactions products with dibutylamine

EC Number:

606-330-7

Molecular formula:

Not specified UVCB - Reaction product of ethoxylated (0-7 EO) propylidenetrimethanol with acrylic acid and dibutylamine (propylidenetrimethanol with acrylic acid : dibutylamine = 5:1)

IUPAC Name:

propylidynetrimethanol, ethoxylated, esters with acrylic acid and its reactions products with dibutylamine

Details on test material:

- Name of test material (as cited in study report): Laromer LR8869
- Physical state: clear colorless liquid
- Analytical purity: The test substance is a complex mixture of isomers and homologous components (see analytical report, study code 11L00263).
- Purity test date: 2012-01-11
- Lot/batch No.: 110009P040
- Expiration date of the lot/batch: 11/2012
- Stability under test conditions: confirmed indirectly by dose formulation analytics
- Storage condition of test material: room temperature, below 25°C, protected from light

Test animals

Species:

mouse

Strain:

NMRI

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Sulzfeld, Germany
- Age at study initiation: 8-9 weeks
- Weight at study initiation: 35.6g (SD +/- 1.5g)
- Assigned to test groups randomly: yes
- Housing: single in Makrolon Type II/III cages with wire mesh top
- Diet (e.g. ad libitum): pelleted standard diet ad lib.
- Water (e.g. ad libitum): tap water ad lib.
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2 °C
- Humidity (%): 45-65%
- Photoperiod (hrs dark / hrs light): 12h/12h

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: corn oil
- Justification for choice of solvent/vehicle: relative non-toxicity
- Amount administered: 10ml/kg b.w.
- Lot/batch no. (if required): MKBF8603V

Duration of treatment / exposure:

24h and 48h (only highest dose tested)

Frequency of treatment:

once

Doses / concentrations

Remarks:

Doses / Concentrations:
500, 1000, 2000mg/kg b.w.
Basis:
actual ingested

No. of animals per sex per dose:

7 (5 for vehicle and positive control)

Control animals:

yes, concurrent vehicle

Positive control(s):

cyclophosphamide
- Route of administration: oral, gavage in sterile water
- Doses / concentrations: 40mg/kg b.w.

Examinations

Tissues and cell types examined:

bone marrow

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: the maximum recommended dose of 2000mg/kg b.w. was tolerated without causing toxic reactions

DETAILS OF SLIDE PREPARATION:
The animals were sacrificed using CO2 followed by bleeding. The femora were removed, the epiphyses were cut off and the marrow was flushed out with foetal calf serum using a syringe. The cell suspension was centrifuged at 1500 rpm (390 x g) for 10 minutes and the supernatant was discarded. A small drop of the re-suspended cell pellet was spread on a slide. The smear was air-dried and then stained with May-Gruenwald (Merck, 64293 Darmstadt, Germany)/Giemsa (Merck, 64293 Darmstadt, Germany). Cover slips were mounted with EUKITT (Kindler, 79110 Freiburg, Germany). At least one slide was made from each bone marrow sample.

METHOD OF ANALYSIS:
Evaluation of the slides was performed using NIKON microscopes with 100x oil immersion objectives. Per animal 2000 polychromatic erythrocytes (PCE) were analysed for micronuclei. To investigate a cytotoxic effect the ratio between polychromatic and normochromatic erythrocytes was determined in the same sample and expressed in polychromatic erythrocytes per 2000 erythrocytes. The analysis was performed with coded slides.

Evaluation criteria:

The study was considered valid as the following criteria are met:
- at least 5 animals per test group can be evaluated.
- PCE to erythrocyte ratio should not be less than 20 % of the vehicle control.
- the positive control shows a statistically significant and biological relevant increase of micronucleated PCEs compared to the vehicle control.

A test item is classified as mutagenic if it induces either a dose-related increase or a clear increase in the number of micronucleated polychromatic erythrocytes in a single dose group.

A test item that fails to produce a biological relevant increase in the number of micronucleated polychromatic erythrocytes is considered non-mutagenic in this system.

Statistics:

nonparametric Mann-Whitney test

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Toxicity:

no effects

Remarks:

only clinical signs evaluated

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): no
- Ratio of PCE/NCE (for Micronucleus assay): not altered
- Statistical evaluation: no significant increase detected

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
The test substance is considered to be non-mutagenic in this micronucleus assay.