Hydrocarbons, C6-C7, n-alkanes, isoalkanes, cyclics, <5% n-hexane

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

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:

September - October 1982

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Test procedure according to national standards

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Reference 2

Reference Type:

publication

Title:

The genetic toxicology of some hydrocarbon and oxygenated solvents

Author:

Brooks, T.M. et al.

Year:

1988

Bibliographic source:

Mutagenesis 3(3): 227-232

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

- limited documentation of cytotoxicity data

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics

EC Number:

920-750-0

IUPAC Name:

hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics

Details on test material:

- Name of test material (as cited in study report): SBP 100/140 (Special Boiling Point Spirit 100/140)
- Substance type: clear, colourless liquid
- Physical state: liquid
- Analytical purity: 100% pure commercial product
- Composition of test material, percentage of components: Special Boiling Point Spirits, SBP 100/140 is a water-white mixture of paraffins and cyclo-paraffins [65% n and isoparaffins, and 35% naphthenes] in the C7-C9 range with 0.01% aromatic hydrocarbons.
- Lot/batch No.: ex Tank 77 P; Indent 9200/9515; Ref LONO42279
- Stability: verified by IR spectra on 14 July 1982 and 20 January 1983; no differences between two spectra
- Storage condition of test material: stored in the dark at ambient temperatures
- Other: Code Number: Q.5811; IP 446/82
- Source: Shell Nederland Raffinaderij B.V., Rotterdam
- test material received on 9 July 1982

Method

Target gene:

S. typhimurium strains: his-operon
E. coli strains: tryptophan-operon

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium, other: TA 1535, TA 1537, TA 1538, TA 98, TA 100

Additional strain / cell type characteristics:

not specified

Species / strain 2

Species / strain / cell type:

E. coli, other: WP2, WP2 uvr A

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

Rat liver homogenate S9 activation, 10 % S9 liver homogenate from Aroclor treated rats

Test concentrations with justification for top dose:

TA 1538 (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
TA 1538 (-S) mix): 0, 15.62, 31.25, 62.5, 125, 250, 500, 1000; (+S9 mix): 0, 31.25, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
TA 1537 (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
TA 1537 (-S9 mix): 0, 3.9, 7.81, 15.62, 31.25, 62.5, 125; (+S9 mix): 0, 31.25, 62.5, 125, 250, 500, 1000, 2000 µg/mL
TA 1535 (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
TA 100 (-S9 mix): 0, 0.31, 0.62, 1.25, 2.5, 5, 10; (+S9 mix): 0, 7.81, 15.62, 31.25, 62.5, 125, 250 µg/mL
TA 100 (-S9 mix): 0, 31.25, 62.5, 125, 250, 500; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
TA 100 (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
TA 98 (-S9 mix): 0, 0.31, 0.62, 1.25, 2.5, 5, 10; (+S9 mix): 0, 7.81, 15.62, 31.25, 62.5, 125, 250 µg/mL
TA 98 (-S9 mix): 0, 3.9, 7.81, 15.62, 31.25, 62.5, 125; (+S9 mix): 0, 31.25, 62.5, 125, 250, 500, 1000, 2000 µg/mL
TA 98 (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
EC WP2 (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL
EC WP2uvrA (-S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000; (+S9 mix): 0, 62.5, 125, 250, 500, 1000, 2000, 4000, 8000 µg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Tween 80 and ethanol

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: TA 1538, TA 98, TA 100: benzo(a)pyrene; TA 1537: neutral red; TA 1535: Sodium azide; and E. coli strains: 4-nitroquinoline-N-oxide

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 30 min
- Incubation period: 48-72 hours

NUMBER OF REPLICATIONS: All test were carried out in triplicate.

Two replicate assays were carried out on different days to confirm reproducibility of results.

Statistics:

none

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 1538, TA 98, TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

Concentrations up to 8000 µg/mL in agar layer cultures did not increase the reverse mutation frequency of S. typhimurium strains either in the presence or absence of a rat liver microsomal activation system.

Cytotoxicity / choice of top concentrations:

other: There was a differential toxicity of the test material to individual strains of Salmonella and the reduction of toxicity in the presence of an S9 mix

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli, other: WP2, WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

Concentrations up to 8000 µg/mL in agar layer cultures did not increase the reverse mutation frequency of E. coli strains either in the presence or absence of a rat liver microsomal activation system

Cytotoxicity / choice of top concentrations:

not specified

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

There was no evidence of test compound precipitation in the assay system.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table: Relative Mutation Rate – Bacterial Experiment I without Metabolic Activation

Concentration	E. coli WP2	E. coli WP2 uvr A	S. typhimurium TA 1535	S. typhimurium TA 1537	S. typhimurium TA 1538	S. typhimurium TA 98	S. typhimurium TA 100
Without S9
3.91	-	-	-	-	-	-	-
7.81	-	-	-	-	-	-	-
15.6	-	-	-	-	-	-	-
31.3	-	-	-	-	-	-	-
62.5	0.8	0.7	1.2	0.1	0.7	0.2	1.4
125	0.9	0.6	0.7	0	0.4	0	0.7
250	0.7	0.8	0.6	0	0	0	0.3
500	0.7	0.7	0.9	0	0	0	0.1
1000	0.8	0.5	0.9	0	0	0	0
2000	0.7	0.6	0.7	0	0	0	0
4000	0.8	0.6	0.6	0	0	0	0
8000	0.6	0.5	0.7	0	0	0	0
Sodium azide 2 µg	-	-	89.8	-	-	-	-
Benzo(a)-pyrene 7 µg	-	-	-	-	1.0	0.7	0.9
4-nitroquinoline-N-oxide 3 µg	39.8	3.2	-	-	-	-	-
Neutral red 7 µg	-	-	-	1.8	-	-	-

Table: Relative Mutation Rate – Bacterial Experiment I with Metabolic Activation

Concentration	E. coli WP2	E. coli WP2 uvr A	S. typhimurium TA 1535	S. typhimurium TA 1537	S. typhimurium TA 1538	S. typhimurium TA 98	S. typhimurium TA 100
Without S9
31.3	-	-	-	-	-	-	-
62.5	0.7	1.3	1.4	0.3	1.5	0.7	1.0
125	0.8	1.0	1.1	0.2	1.3	1.3	0.8
250	0.9	1.0	0.8	0.6	1.5	1.2	1.0
500	0.7	0.8	1.0	0.2	1.0	0.7	1.3
1000	0.7	0.7	1.0	0.4	0.7	0.2	1.4
2000	1.1	0.9	1.1	0	0.4	0	1.2
4000	1.0	1.0	1.0	0	0.7	0	1.1
8000	0.8	1.0	0.5	0	0.7	0	1.1
Sodium azide 2 µg	-	-	15.2	-	-	-	-
Benzo(a)-pyrene 7 µg	-	-	-	-	3.7	5.3	3.1
4-nitroquinoline-N-oxide 3 µg	1.2	3.6	-	-	-	-	-
Neutral red 7 µg	-	-	-	4.4	-	-	-

Table: Relative Mutation Rate – Bacterial Experiment II without Metabolic Activation

Concentration	E. coli WP2	E. coli WP2 uvr A	S. typhimurium TA 1535	S. typhimurium TA 1537	S. typhimurium TA 1538	S. typhimurium TA 98	S. typhimurium TA 100
Without S9
3.91	-	-	-	1.1	-	0.8	-
7.81	-	-	-	0.6	-	0.9	-
15.6	-	-	-	0.7	0.9	0.8	-
31.3	-	-	-	0.4	1.0	0.8	1.1
62.5	1.0	1.0	1.2	0.4	0.7	0.4	1.0
125	1.0	1.0	0.9	0	0.5	0	0.9
250	0.7	0.8	1.4	-	0.2	-	1.0
500	0.9	0.7	1.1	-	0	-	0.7
1000	1.0	0.8	0.9	-	0	-	-
2000	0.7	0.9	0.7	-	-	-	-
4000	0.9	0.8	0.9	-	-	-	-
8000	0.8	0.7	0.9	-	-	-	-
Sodium azide 2 µg	-	-	86.6	-	-	-	-
Benzo(a)-pyrene 7 µg	-	-	-	-	1.1	1.1	1.3
4-nitroquinoline-N-oxide 3 µg	36.7	5.3	-	-	-	-	-
Neutral red 7 µg	-	-	-	1.2	-	-	-

Table: Relative Mutation Rate – Bacterial Experiment II with Metabolic Activation

Concentration	E. coli WP2	E. coli WP2 uvr A	S. typhimurium TA 1535	S. typhimurium TA 1537	S. typhimurium TA 1538	S. typhimurium TA 98	S. typhimurium TA 100
Without S9
31.3	-	-	-	1.1	0.9	0.8	-
62.5	0.9	0.9	0.7	1.2	1.1	1.0	1.1
125	1.1	1.0	1.2	1.1	1.1	0.9	1.0
250	1.4	0.8	0.8	1.4	1.1	0.8	1.1
500	1.2	1.0	0.9	1.0	1.1	0.8	1.2
1000	1.0	1.1	0.8	0.5	0.9	0.4	0.9
2000	0.8	1.0	1.1	0	0.9	0.1	0.9
4000	0.8	0.9	0.8	-	0.5	-	0.9
8000	1.0	0.7	0.7	-	0.4	-	0.9
Sodium azide 2 µg	-	-	18.2	-	-	-	-
Benzo(a)-pyrene 7 µg	-	-	-	-	9.7	6.1	6.5
4-nitroquinoline-N-oxide 3 µg	1.1	2.8	-	-	-	-	-
Neutral red 7 µg	-	-	-	5.2	-	-	-

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

The purpose of this study was to determine the mutagenicity of the test substance hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics. An Ames reverse mutation assay was done on S. typhimurium strains, TA 1535, TA 1537, TA 1538, TA 98 and TA 100, and E. coli strains WP2 uvr A and WP2, both with and without metabolic activation. Species were tested at concentrations ranging from 0.31 to 8000 ug/plate. The test plates were incubated for 48 -72 hrs, and then the number of colonies were counted. For all species, there was no significant increase in the number of revertants as compared to negative controls. Positive controls showed a significant increase in mutations. The test substance is not mutagenic.

Executive summary:

The purpose of this study was to determine the mutagenicity of the test substance hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics. An Ames reverse mutation assay was done on S. typhimurium strains, TA 1535, TA 1537, TA 1538, TA 98 and TA 100, and E. coli strains WP2 uvr A and WP2, both with and without metabolic activation. Species were tested at concentrations ranging from 0.31 to 8000 ug/plate. The test plates were incubated for 48 -72 hrs, and then the number of colonies were counted. For all species, there was no significant increase in the number of revertants as compared to negative controls. Positive controls showed a significant increase in mutations. The test substance is not mutagenic.