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Diss Factsheets

Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
1985-11-26 to 1986-02-05
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study is classified as reliable without restriction because it was conducted according to or similar to guideline study OECD TG 476.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1986
Report date:
1986
Reference Type:
publication
Title:
Laboratory procedure for assessing specific locus mutations at the TK locus in cultured L5178Y mouse lymphoma cells
Author:
Clive, D. and Spector, J.F.S.
Year:
1975
Bibliographic source:
Mutation Res., vol. 31, pp. 17-29

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay

Test material

Constituent 1
Reference substance name:
64742-05-8
IUPAC Name:
64742-05-8
Constituent 2
Reference substance name:
Light paraffinic distillate solvent extract
IUPAC Name:
Light paraffinic distillate solvent extract
Test material form:
liquid: viscous
Details on test material:
- Name of test material (as cited in study report): API 83-16
- Substance type: Petroleum
- Physical state: Clear dark brown liquid
- Viscosity, SSU: 67.2 at 100°F
- API Gravity: 16.7
- Flash Point: (°F) 335
- Pour Point (°F): 20
- Aniline Point (°F): 95.5
- Sulfur, Wt %: 2.64

- Composition of test material ASTM D-2007, Wt. %:
Saturates: 29.2
Aromatics 68.6

Method

Species / strain
Species / strain / cell type:
mouse lymphoma L5178Y cells
Details on mammalian cell type (if applicable):
- Type and identity of media: RPMI-1640 medium supplemented with pluronic solution, L-glutamine, sodium pyruvate, antibiotics, and horse serum (10% by volume)
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes/no No data
- Periodically "cleansed" against high spontaneous background: yes/no No data
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Test concentrations with justification for top dose:
25, 50, 75, 100, 150 and 200 µL/mL without activation and at 12.5, 25, 50, 75, 100 and 150 µL/mL with activation
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Ethanol
- Justification for choice of solvent/vehicle: More compatible with cell viability than acetone.
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
ethanol
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
Migrated to IUCLID6: for nonactivation studies
Details on test system and experimental conditions:
METHOD OF APPLICATION: In medium


DURATION
- Preincubation period: 10 to 14 days
- Expression time (cells in growth medium): 2 days



SELECTION AGENT (mutation assays): 3µg/mL 5-trifluorothymidine (TFT)

NUMBER OF REPLICATIONS: not reported


NUMBER OF CELLS EVALUATED: not reported

Evaluation criteria:
The minimum criterion considered necessary to demonstrate mutagenesis for any given treatment will be a mutant frequency that is at least 150% of the concurrent background frequency plus 10 x10 e-6. The background frequency is defined as the average mutant frequency of the solvent negative controls.

Results and discussion

Test results
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
not determined
Vehicle controls validity:
not examined
Untreated negative controls validity:
not examined
Positive controls validity:
not examined
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):
positive with metabolic activation
positive without metabolic activation

In the absence of metabolic activation, a dose-response relationship was observed from 75 µL/mL to 200 µL/mL DAE with decreasing relative growth percentages and increasing mutant frequency (in 10E-6 units). The minimum criterion for mutagenesis in this assay was a mutant frequency exceeding 67.9x10-6. Concentrations of 150 µL/mL and 200 µL/mL induced mutant frequencies that exceeded the minimum criterion. Hence the tested material was considered mutagenic without activation.

In the presence of metabolic activation, a dose-response relationship was observed from 75 µL/mL to 150 µL/mL DAE with decreasing relative growth percentages and increasing mutant frequency (in 10E-6 units). The minimum criterion for mutagenesis in this assay was a mutant frequency exceeding 73.5x10-6. Treatments from 25 µL/mL to 150 µL/mL induced increases in the mutant frequency, which ranged from 1.8-fold to 3.2-fold above the minimum criterion. There was a general trend toward higher mutant frequencies at higher concentrations of the test material. The tested material was therefore considered mutagenic with activation.

Light paraffinic distillate solvent extract is therefore mutagenic with and without activation under the conditions specified in this assay.
Executive summary:

In a mammalian cell gene mutation assay mouse lymphoma L5178Y cells cultured in vitro were exposed to light paraffinic distillate solvent extract in ethanol at concentrations of 25, 50, 75, 100, 150 and 200 µL/mL without activation and at 12.5, 25, 50, 75, 100 and 150 µL/mL with activation.

In the absence of metabolic activation, a dose-response relationship was observed from 75 µL/mL to 200 µL/mL DAE with decreasing relative growth percentages (i.e., 119.7% to 2.2%) and increasing mutant frequency (in 10E-6 units). The minimum criterion for mutagenesis in this assay was a mutant frequency exceeding 67.9x10-6. Concentrations of 150 µL/mL and 200 µL/mL induced mutant frequencies that exceeded the minimum criterion. Hence the tested material was considered mutagenic without activation. In the presence of metabolic activation, a dose-response relationship was observed from 75 µL/mL to 150 µL/mL DAE with decreasing relative growth percentages and increasing mutant frequency (in 10E-6 units). The minimum criterion for mutagenesis in this assay was a mutant frequency exceeding 73.5x10-6. Treatments from 25 mL/mL to 150 mL/mL induced increases in the mutant frequency, which ranged from 1.8-fold to 3.2-fold above the minimum criterion.

There was a general trend toward higher mutant frequencies at higher concentrations of the test material. The tested material was therefore considered mutagenic with activation. Light paraffinic distillate solvent extract is therefore mutagenic with and without activation under the conditions specified in this assay.

This study received a Klimisch score of one and is considered reliable without restriction because it was conducted according to or similar to guideline study OECD TG 476.