Hydrocarbons, C7-C8, isoalkanes, <2% aromatics

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

There is no data available for Hydrocarbons, C7-C8, isoalkanes, <2% aromatics. However, data is available for structural analogues, Hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics and 2,2,4-trimethylpentane, andpresented in the dossier. This data is read across to based on analogue read across and a discussion and report on the read across strategy is provided as an attachment in IUCLID Section 13.

The available data indicate that hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics and 2,2,4-trimethylpentane are not genotoxic.

Negative Ames test with S. typhimurium TA 1535, TA 1537, TA 1538, TA 98 and TA 100, and E. coli WP2 and WP2 uvr A, with and without metabolic activation.

Negative results in mammalian chromosomal aberration and gene mutation tests, the latter with and without metabolic activation.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

read-across from supporting substance (structural analogue or surrogate)

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

Justification for type of information:

A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across: supporting information

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

Remarks:

Not specified

Type of assay:

bacterial reverse mutation assay

Target gene:

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

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 / 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

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

Key result

Species / strain:

S. typhimurium TA 1535

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

Key result

Species / strain:

S. typhimurium TA 1537

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

Key result

Species / strain:

S. typhimurium TA 1538

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

Key result

Species / strain:

S. typhimurium TA 98

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

Key result

Species / strain:

S. typhimurium 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

Key result

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'.

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	-	-	-

Conclusions:

Interpretation of results:
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 µg/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.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

read-across from supporting substance (structural analogue or surrogate)

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.

Justification for type of information:

A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across: supporting information

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

GLP compliance:

no

Remarks:

Not specified

Type of assay:

in vitro mammalian chromosome aberration test

Species / strain / cell type:

mammalian cell line, other: Rat liver epithelial-type line (RL4) cells

Details on mammalian cell type (if applicable):

- Type and identity of media: Stock culture media and assay media was Minimal Essential Medium supplemented with 10% fetal calf serum and 1% non-essential amino acids.

Additional strain / cell type characteristics:

other: metabolic competent

Metabolic activation:

without

Metabolic activation system:

RL4 cells are metabolically competent.

Test concentrations with justification for top dose:

0, 2.5, 5.0, and 10 mg/mL

Vehicle / solvent:

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

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

7,12-dimethylbenzanthracene

Details on test system and experimental conditions:

see "Any other information on materials and methods"

Key result

Species / strain:

mammalian cell line, other: RL4 cells, metabolic competent

Metabolic activation:

not applicable

Genotoxicity:

negative

Remarks:

There was no significant increase in chromosome damage

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Exposure of RL4 cells to 20 µg/mL test substance resulted in inhibition of cell proliferation by 50 %. Therefore final test concentrations of 2.5, 5, and 10 µg/mL were chosen

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

A maximum dose level of 10 µg/mL was determined in the chromosome assay. There was no significant increase beyond control limits in the frequency of chromatid breaks or total chromatid aberrations in rat liver cells exposed to the test material up to 10 µg/mL indicating that the compound did not induce chromosome damage in rat liver cells. A small increase in the percentage of cells showing gaps in exposed cultures was well within historical control ranges. Cultures exposed to the positive control material, DMBA, showed an increase in the frequency of chromosome damage.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Table: Metaphase chromosome analysis of RL₄cells

	0 µg/ml	2.5 µg/ml	5.0 µg/ml	10.0 µg/ml	1.0 µg/ml 7,12-dimethylbenzanthracene
Polyploidy (% cells)	1.3	1.3	1.3	1.3	0.5
Chromatid gaps (% cells)	0.3	0.7	0.9	1.0	5.5
Multiple aberrations (% cells)	-	-	-	-	-
Severe damage (% cells)	-	-	-	-	-
Chromatid aberrations (% cells)	0.7	-	0.9	-	4.0
Chromosome aberrations (% cells)	-	-	-	-	1.5
Chromatid gaps (frequency per cell)	0.003	0.007	0.009	0.010	0.060
Chromatid breaks (frequency per cell)	0.007	-	0.004	-	-
Chromatid exchange (frequency per cell)	-	-	0.004	-	-
Chromosome breaks (frequency per cell)	-	-	-	-	0.015
Chromosome exchange (frequency per cell)	-	-	-	-	-

Conclusions:

Interpretation of results: negative

This study examined the potential for the test substance, hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics, to cause chromosomal aberrations in rat liver RL4 cells. Cells were exposed to concentrations of 0, 2.5, 5.0, and 10 ug/mL of test substance, and then examined for chromosomal aberrations including polyploidy, chromatid gaps, and chromatid exchanges. The positive control substance was 7,12 -dimethylbenzanthracene. No significant increase in the frequency of chromosomal damage was observed. Based on the study design the test substance showed no genetic toxicity.

Executive summary:

This study examined the potential for the test substance, hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics, to cause chromosomal aberrations in rat liver RL4 cells. Cells were exposed to concentrations of 0, 2.5, 5.0, and 10 µg/mL of test substance, and then examined for chromosomal aberrations including polyploidy, chromatid gaps, and chromatid exchanges. The positive control substance was 7,12 -dimethylbenzanthracene.

 

No significant increase in the frequency of chromosomal damage was observed. Based on the study design the test substance showed no genetic toxicity.

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well documented publication which meets basic scientific principles

Justification for type of information:

A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across: supporting information

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

yes

Remarks:

- using microtiter plates

GLP compliance:

not specified

Type of assay:

mammalian cell gene mutation assay

Target gene:

thymidine kinase

Species / strain / cell type:

human lymphoblastoid cells (TK6)

Details on mammalian cell type (if applicable):

- Type and identity of media: RPMI 1640 medium supplemented with 15% heat-inactivated horse serum

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

with Aroclor induced-rat liver homogenate

Test concentrations with justification for top dose:

5 % v/v TMP in DMEM medium, administered undiluted or as 50 % (1:1 saturated to normal medium)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMEM medium

Untreated negative controls:

yes

Remarks:

with and without S9

Negative solvent / vehicle controls:

yes

Remarks:

DMEM medium

Positive controls:

yes

Remarks:

for S9 activated cultures

Positive control substance:

benzo(a)pyrene

Remarks:

Migrated to IUCLID6: 15 µM

Untreated negative controls:

yes

Remarks:

with and without S9

Negative solvent / vehicle controls:

yes

Remarks:

DMEM medium

Positive controls:

yes

Remarks:

for non-activated cultures

Positive control substance:

ethylmethanesulphonate

Remarks:

Migrated to IUCLID6: 0.2 mM

Details on test system and experimental conditions:

see "any other information on materials and methods"

Key result

Species / strain:

human lymphoblastoid cells (TK6)

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

TMP did not induce significant increases in the mutation frequency at the thymidine kinase locus.

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Cell survival in TMP-saturated medium with and without metabolic activation was greater than 50-60 %.

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

The elevated mutation frequencies of positive control compounds were as expected.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
Cell survival in positive control treated cultures was 40 or 30% for benzo(a)pyrene and EMS, respectively

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Conclusions:

Interpretation of results: negative

Based on the study design there is no incidence of increased genetic toxicity caused by the test substance.

Executive summary:

Based on the study design there is no incidence of increased genetic toxicity caused by the test substance.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

There is no data available for Hydrocarbons, C7-C8, isoalkanes, <2% aromatics. However, data is available for structural analogue, Hydrocarbons, C7-C9, isoalkanes, <2% aromatics and presented in the dossier. This data is read across to based on analogue read across and a discussion and report on the read across strategy is provided as an attachment in IUCLID Section 13.

Hydrocarbons C7-C9, isoalkanes, <2% aromatics

Negative in dominant lethal, micronucleus and unscheduled DNA synthesis assays.

Link to relevant study records

Reference

Endpoint:

in vivo mammalian germ cell study: cytogenicity / chromosome aberration

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

July 1978 - January 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline with acceptable restrictions.

Justification for type of information:

A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across: supporting information

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 478 (Genetic Toxicology: Rodent Dominant Lethal Test)

Principles of method if other than guideline:

Similar to OECD 478, only two dose levels were used.

GLP compliance:

not specified

Type of assay:

rodent dominant lethal assay

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Wilmington, Massachusetts
- Age at study initiation: males: 10 weeks at initiation of pre-treatment mating period; females: 9 weeks at initiation of week 1 of pre-treatment mating period; females: 8 weeks at initiation of week 1 of post-treatment mating period
- Housing: males: individually housing during the treatment period; females: individually housing during the pre- and post-mating period
- Diet: Purina Laboratory Chow; ad libitum; except chamber-exposed groups during the treatment period
- Water: ad libitum; except chamber-exposed groups during the treatment period

Route of administration:

inhalation

Vehicle:

none

Details on exposure:

TYPE OF INHALATION EXPOSURE: whole body


GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: The stainless steel and glass chambers had a total volume of one cubic metre with an effective volume of 760 L.
- Air flow rate: 132 L/min
- Air change rate: every 7.6 min
- The test substance was placed in fritted-bottom, gas-washing bottles. Dry air, at various flow rates, was passed through the test material and the vapour-air mixtures entered the chamber air inlets and were further diluted with room air to provide the desired concentrations.


TEST ATMOSPHERE
Atmospheric sampling was performed using a Wilks Scientific Corp., Miran IA Ambient Air Analyzer (long pathlength infrared). On each exposure day three samples were drawn from each exposure chamber (at 1, 3 and 5 hours) and the exposure concentrations calculated by comparing the absorption of this sample to the standard curve.

Duration of treatment / exposure:

8 consecutive weeks

Frequency of treatment:

6 hours/day, 5 days/week

Post exposure period:

4 weeks

Remarks:

Doses / Concentrations:
400 and 1200 ppm
Basis:
nominal conc.

No. of animals per sex per dose:

only males were exposed: 10/group

Control animals:

yes, sham-exposed

Positive control(s):

triethylenemelamine
- Route of administration: intraperitoneally on a single day, at least 2 hours prior to mating
- Doses level/ volume: 0.5 mg/kg body weight; 1 mL/kg body weight
- Vehicle: 0.9 % injectable normal saline solution

Tissues and cell types examined:

Males: testes, seminal vesicles, epididymides, prostate tissues
Females: uteri

Details of tissue and slide preparation:

METHOD OF ANALYSIS:
Males: Seminal vesicles, epididymides, prostate and any abnormal lesion or tissue masses were preserved in 10 % neutral buffered formalin. Testes were stored for 48 hours in Bouin´s solution and subsequently placed in formalin. Tissues from 5 randomly chosen males from each group were examined.
Females: The uteri from all females (pregnant and non-pregnant) were appropriately identified and preserved intact in a 10 % neutral buffered formalin solution.

Statistics:

Comparisons were made during the treatment and post-treatment periods between negative control, positive control and test substance-treated groups by the Chi-square test. Absolute data were compared by using the F-test and Student´s t-test. When variances differed significantly, Student´s t-test was appropriately modified using Cochran´s approximation (t´). Corpora lutea and implantation data were evaluated by the one-tailed test.

Key result

Sex:

male/female

Genotoxicity:

negative

Remarks:

At dose level of 400 and 1200 ppm the test substance was not mutagenic in rats by the dominant-lethal test.

Toxicity:

no effects

Vehicle controls validity:

other: no vehicle was used

Negative controls validity:

valid

Positive controls validity:

valid

No treatment-related effects were observed for mortality, in-life physical or necropsy examinations and for body weights in males.

No treatment effect was indicated from microscopic evaluation of testes, seminal vesicles, epididymides and prostate tissues from 5 randomly chosen males from each exposure group.

In one animal of the 400 ppm exposure group a caseous mass was observed in the left epididymis. This caseous mass was seen microscopically to be a spermatic granuloma. Since this was observed in a single animal, this is not considered to be treatment related. Several animals (one in control, one in 100 ppm group, one in 1200 ppm group) presented a purulent prostatitis which was of probable bacterial etiology.

Pregnancy rates, implantation data and implantation efficiency values and fetal death data for females mated to treated males were considered comparable to data for females mated to negative control males for each week of the post-treatment mating period. At dose level of 400 and 1200 ppm the test substance was not mutagenic in rats by the dominant-lethal test.

Positive control:

Females mated to the positive control males had fewer implants and lower implantation efficiency values (indicative of pre-implantation loss) than females mated to the negative control males. Fetal death parameters (mean and expressed as a percentage of total implants) for these same females were significantly increased during the post-treatment mating interval.

Conclusions:

Interpretation of results: negative
Under the conditions of the test no negative effects were induced by the test substance, hydrocarbons, C7-C9, isoalkanes.

Executive summary:

Under the conditions of the test no negative effects were induced by the test substance, hydrocarbons, C7-C9, isoalkanes.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

There is no data available for Hydrocarbons, C7-C8, isoalkanes, <2% aromatics. However, data is available for structural analogues, Hydrocarbons, C7-C9, isoalkanes, <2% aromatics; Hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics; and 2,2,4-trimethylpentane, and presented in the dossier. This data is read across to based on analogue read across and a discussion and report on the read across strategy is provided as an attachment in IUCLID Section 13.

In Vitro

In vitro Gene Mutation in Bacteria

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

A reliable bacterial reverse mutation assay (Ames test) was conducted with hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics following a protocol similar to OECD 471. The pre-incubation procedure was performed with Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100, and Escherichia coli strains WP2 uvr A and WP2. The strains were exposed to concentrations ranging from 0.31 to 8000 µg/plate for 30 min both with and without metabolic activation prior to plating and further incubation for 48 -72 h.For all strains tested, there was no significant increase in the number of revertants as compared to negative controls. An initial cytotoxicity assay with S. typhimurium TA100 showed that the test substance was extremely cytotoxic, especially in the absence of S9 fraction. Therefore, initial mutation assays were conducted over a low dose range (0.31 -10 µg per mL) with strains TA100 and TA98. Virtually no cytotoxicity was observed at these concentrations, and so more appropriate concentrations were tested for all strains in subsequent assays. Salmonella strains were more sensitive to the cytotoxic effect of the test substance than E. coli strains, and cytotoxicity was greater in the absence of S9-fraction. In conclusion, the test substance was not mutagenic in this assay (Shell Chemicals, 1983a; Brooks et al., 1988).

In Vitro Chromosome Aberration in Mammalian Cells

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

The potential of hydrocarbons, C7-C9, n-alkanes, isoalkanes, cyclics to cause chromosomal aberrations in rat liver RL4 cells was tested with a method comparable to OECD 473. Cells were exposed to concentrations of 0, 2.5, 5, and 10 µg/mL of test substance for 22 h, and then examined for chromosomal aberrations including polyploidy, chromatid gaps, and chromatid exchanges. Exposure of RL4 cells to 20 µg/mL test substance resulted in inhibition of cell proliferation by 50%. Therefore 10 µg/mL was chosen as the maximum dose level. No significant increase in the frequency of chromosomal damage was observed. Under the conditions of this study, the test material was not clastogenic (Shell Chemicals, 1983b; Brooks et al., 1988).

In Vitro Gene Mutation in Mammalian Cells

Iso-octane (CAS No. 540-84-1) was tested in a mammalian cell gene mutation assay performed according to OECD 476. The test material was prepared by adding iso-octane at a final concentration of 5 % v/v in culture (DMEM) medium and stirred overnight at room temperature in a foil wrapped, capped parafilm-sealed bottle to saturate the medium. Human lymphoblastoid cells (TK6) were exposed to 100 or 50 % of this saturated DMEM medium with and without metabolic activation for 3 h and allowed for expression for 4 to 8 days. Both with and without metabolic activation, iso-octane did not induce significant increases in the mutation frequency at the thymidine kinase locus and cell survival in iso-octane-saturated medium was greater than 50-60 %. Based on the study design there was no incidence of increased genetic toxicity caused by the test substance (Richardson et al., 1986).

 

In vivo

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

Hydrocarbons, C7-C9, isoalkanes tested in a dominant lethal study (similar to OECD 478) showed no evidence of genotoxicity in the germ cells of treated male rats exposed to 400 or 1200 ppm by inhalation (ExxonMobil Chemical,1980a).

Based on the analogue approach, these results suggest that hydrocarbons, C7-C8, isoalkanes, <2% aromatics is not expected to induce genotoxicity in vivo.

Justification for classification or non-classification

Based on available read across data, Hydrocarbons, C7-C8, isoalkanes, <2% aromatics does not warrant classification for genetic toxicity under the Regulation (EC) 1272/2008 on classification, labelling and packaging of substances and mixtures (CLP).