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Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
18 Jan to 12 Feb 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD guideline study to GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1999
Report date:
1999

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
1-(N,N-bis(2-hydroxyethyl)amino)propan-2-ol
EC Number:
229-764-5
EC Name:
1-(N,N-bis(2-hydroxyethyl)amino)propan-2-ol
Cas Number:
6712-98-7
Molecular formula:
C7H17NO3
IUPAC Name:
1-[bis(2-hydroxyethyl)amino]propan-2-ol
Test material form:
other: Transparent, colourless viscous liquid
Details on test material:
- Name of test material (as cited in study report): Diethanolisopropanolamine (DEIPA)
- Substance type: Technical product
- Physical state: Liquid
- Analytical purity: 93%
- Impurities (identity and concentrations): No data
- Composition of test material, percentage of components: No data
- Isomers composition: No data
- Purity test date: Not specified
- Lot/batch No.: 980035B
- Expiration date of the lot/batch: Not specified
- Stability under test conditions: Not specified
- Storage condition of test material: Not specified

Method

Target gene:
S. typhimurium: Histidine locus
E. coli WP2 uvr A: Tryptophan locus
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
S9 fraction from Aroclor-induced rat liver
Test concentrations with justification for top dose:
100, 330, 1000, 3330, 5000 μg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: Not specified [OECD guideline recommends the use of aqueous solvent/vehicle]
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
2-nitrofluorene
sodium azide
benzo(a)pyrene
other: 2-aminoanthracene and ICR-191
Details on test system and experimental conditions:
METHOD OF APPLICATION: preincubation

Preincubated in the presence of the tester strain for 20 mins at 37oC before adding the overlay agar and pouring onto the surface of minimal agar plates. The plates were incubated for 52 +/- 4 hours at 37 +/- 2oC.

DURATION
- Preincubation period:20 +/- 2 minutes
- Exposure duration: 48-56 hours

SELECTION AGENT (mutation assays): Deficiency in histidine or tryphtophan for selection of non-amino acid requiring mutants.

NUMBER OF REPLICATIONS: 2 independant assays

DETERMINATION OF CYTOTOXICITY
- Method: A decrease in the number of revertant colonies per plate and/or by a thinning or disappearance of the bacterial background lawn indicates cytotoxicity
Evaluation criteria:
The test article had to produce at least a 3-fold (TA98, TA1535, TA1537 and WP2uvr A) or 2-fold (TA100) dose related and reproducible increase in the mean revertants per plate of at least one tester strain over the mean revertants per plate of the appropriate vehicle control
Statistics:
None reported

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
Not applicable

RANGE-FINDING/SCREENING STUDIES: Ten doses of test article from 6.67 to 5000 ul per plate were tested. No cytotoxicity observed with TA100 and WP2uvrA either in presence or absence of metabolic activation system, normal backgroud lawn evident and no decrease in number of revertants per plate. No test article precipitate observed at any of the doses tested.

COMPARISON WITH HISTORICAL CONTROL DATA: Revertants were within the range of historical data for the vehicle and positive controls with each of the strains tested, both with and without S9.

ADDITIONAL INFORMATION ON CYTOTOXICITY: None reported
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

In a GLP study, conducted according to OECD guideline 471, DEIPA showed no convincing evidence of mutagenic activity when tested at concentrations of up to 5000 µg/plate in a bacterial reverse mutation assay with Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and Escherichia coli strain WP2uvrA, with and without metabolic activation (S9).
Executive summary:

In a GLP study, conducted according to OECD guideline 471, DEIPA was assessed for mutagenicity in a bacterial reverse mutation assay.

Two independent studies using the preincubation method were performed using Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and Escherichia coli strain WP2uvrA in the presence and absence of a rat liver metabolic activation (S9 mix). Concentrations of 100, 333, 1000, 3330, and 5000 µg/plate of DEIPA were tested, along with concurrent positive and vehicle controls, all plated in triplicate.The positive control substances all showed mutagenic activity, demonstrating that the test was valid.

Following incubation of the test material, tester strain and S9 mix (or phosphate buffer), the number of revertant colonies per plate was counted. Cytotoxicity was evaluated based on the condition of the bacterial background lawn. No precipitation of the test article was obseved at any dose level.

Under the conditions of this study, DEIPA did not cause a positive increase in the number of revertant colonies per plate and no cytotoxicity was evident, with any of the tester strains, at any concentration, either in the presence or absence S9.

DEIPA showed no convincing evidence of mutagenic activity.

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