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Diss Factsheets

Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro DNA damage and/or repair study
Remarks:
Type of genotoxicity: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from USEPA HPVreport

Data source

Reference
Reference Type:
review article or handbook
Title:
Genetic Toxicity in vitro study for Ethanol, 2-(hydroxymethylamino) _ (CAS# 34375-28-5)
Author:
US EPA HPV Chemical Challenge Program- Troy Corporation
Year:
2005
Bibliographic source:
U.S Environmental Protection Agency/ High Production Volume Information System (HPVIS) 2005

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: EPA Guideline 84-2(b)
Principles of method if other than guideline:
Genetic toxicity study in vitro was conducted to assess the ability of the Ethanol, 2-(hydroxymethylamino) to induce unscheduled DNA synthesis in primary cultures of adult rat hepatocytes.
GLP compliance:
no
Type of assay:
other: unscheduled DNA Synthesis

Test material

Constituent 1
Chemical structure
Reference substance name:
2-(hydroxymethylamino)ethanol
EC Number:
251-974-0
EC Name:
2-(hydroxymethylamino)ethanol
Cas Number:
34375-28-5
Molecular formula:
C3H9NO2
IUPAC Name:
2-[hydroxy(methyl)amino]ethanol
Details on test material:
Details on test material
- Name of test material (as cited in study report): Ethanol, 2-(hydroxymethylamino)
- Molecular formula (if other than submission substance): C3-H9-N-O2
- Molecular weight (if other than submission substance): 91.1091 g/mole
- Substance type: Organic
- Physical state: Liquid
Purity: 100 % (estimated by calculation)
- Impurities (identity and concentrations): No data available
Specific details on test material used for the study:
- Name of test material: Troysan 174, (2[(Hydroxymethyl)amino] ethanol)
- IUPAC name: 2-[hydroxy(methyl)amino]ethanol
- Molecular formula: C3H9NO2
- Molecular weight: 91.1091 g/mol
- Substance type: Organic
- Physical state: No data
- Purity: 100 %
- Impurities (identity and concentrations): No data

Method

Target gene:
No data
Species / strain
Species / strain / cell type:
hepatocytes: Rat/ Fischer 344/ adult hepatocytes -
Details on mammalian cell type (if applicable):
No data
Additional strain / cell type characteristics:
not specified
Cytokinesis block (if used):
No data
Metabolic activation:
not specified
Metabolic activation system:
No data
Test concentrations with justification for top dose:
0.78125, 1.5625, 3.125, 6.25, 12.5, 25.0, 50.0 and 100.0 ug/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The chemical was soluble in DMSO
Controls
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
2-acetylaminofluorene
other: Michler's ketone (direct acting) at 2.0 and 8.0 ug/ml
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium
- Cell density at seeding (if applicable): No data

DURATION
- Preincubation period: No data
- Exposure duration: No data
- Expression time (cells in growth medium): No data
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data

SPINDLE INHIBITOR (cytogenetic assays): No data

STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: Duplicate

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED:

NUMBER OF CELLS EVALUATED: No data

NUMBER OF METAPHASE SPREADS ANALYSED PER DOSE (if in vitro cytogenicity study in mammalian cells): No data

CRITERIA FOR MICRONUCLEUS IDENTIFICATION: No data

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data
- Any supplementary information relevant to cytotoxicity: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Methods, such as kinetochore antibody binding, to characterize whether micronuclei contain whole or fragmented chromosomes (if applicable): No data

- OTHER: This study was conducted to assess the ability of the test compound to induce
unscheduled DNA synthesis in primary cultures of adult rat hepatocytes, as measured by silver grain counts in photographic emulsion formed by radiation from [6 3H]-thymidine taken up by the cells. The cells were examined microscopically at approximately 1000x magnification under oil immersion using a Leitz Dialux 20L microscope.
Rationale for test conditions:
No data
Evaluation criteria:
Unscheduled DNA Synthesis was measured by counting nuclear grains and subtracting the average number of cytoplasmic grains in 3 nuclear sized areas
adjacent to each nucleus (background count). This value was referred to as the net nuclear grain count. The cell cultures were scored, and the results assessed according to the criteria of Butterworth et al, in conjunction with historical data generated in-house
Statistics:
No data available

Results and discussion

Test results
Species / strain:
hepatocytes: Rat/ Fischer 344/ adult hepatocytes -
Metabolic activation:
not specified
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
50 – 100 ug/ml
Vehicle controls validity:
valid
Remarks:
Vehicle controls were considered valid, having met the requirements as specified by the acceptance criteria
Untreated negative controls validity:
not specified
Positive controls validity:
valid
Remarks:
Both direct and indirect acting positive controls demonstrated the sensitivity of the assays.
Additional information on results:
No data

Applicant's summary and conclusion

Conclusions:
Unscheduled DNA Synthesis was not demonstrated by 2-[(Hydroxymethyl) amino] ethanol when tested in dimethylsulphoxide at concentrations extending into the toxic range. Therefore, the substance 2-[(Hydroxymethyl) amino] ethanol is non genotoxic to adult rat hepatocytes.
Executive summary:

Genetic toxicity study in vitro was conducted to assess the ability of the Ethanol, 2-(hydroxymethylamino) to induce unscheduled DNA synthesis in primary cultures of adult rat hepatocytes. Cultures were established with cells derived from the collagenase-perfused liver of a young adult male Fischer 344 rat. The culture was exposed to Ethanol, 2-(hydroxymethylamino) at concentrations of 0.78125, 1.5625, 3.125, 6.25, 12.5, 25.0, 50.0 and 100.0 ug/ml in 2 independent experiments. Positive control was used Michler's ketone (direct acting) and 2-Acetylamino fluorine (2-AAF) (indirect acting). The cells were examined microscopically at approximately 1000x magnification under oil immersion using a Leitz Dialux 20L microscope. Unscheduled DNA Synthesis was measured by counting nuclear grains and subtracting the average number of cytoplasmic grains in 3 nuclear sized areas adjacent to each nucleus (background count). This value was referred to as the net nuclear grain count. In the first assay, there were no evidence of of unscheduled DNA synthesis up to 25 μg/ml. In the second assay, results were obtained up to 25 µg /ml with no evidence of unscheduled DNA synthesis at concentrations of 12.5 µg /ml and lower. Cell toxicity was observed at concentrations of 50 and 100 µg /ml. At the concentration of 25.5 µg/ml there was, however, a slight increase in the mean net grains per nucleus, accompanied by an increase in the percentage of cells adjudged to be in repair. The increase obtained did not meet the criteria required for a positive response, was not similarly observed in the first experiment and was not associated with a dose-related trend at lower concentrations. Unscheduled DNA Synthesis was not demonstrated by 2-[(Hydroxymethyl) amino] ethanol when tested in dimethylsulphoxide at concentrations extending into the toxic range. Therefore, the substance 2-[(Hydroxymethyl) amino] ethanol is non genotoxic to adult rat hepatocytes.