Hydrocarbons, C10-C13, n-alkanes

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

AMES data is available for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics. Additional in vitro genetic toxicity data is available for structural analogues Hydrocarbons, C10-C13, isoalkanes, <2% aromatics and Hydrocarbons, C11 -C14, n-alkanes, isoalkanes, cyclics, <2% aromatics. This data is read across to Hydrocarbons, C10 -C13, n-alkanes, <2% aromatics 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 genetic toxicity test listed below had a negative result for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics.

Genetic Toxicity in vitro – Bacterial reverse mutation assay (OECD 471)

 

All read across genetic toxicity tests listed below had negative results for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics.

 

Genetic Toxicity in vitro – Mammalian Chromosome Aberration Test (OECD TG 473)

Genetic Toxicity in vitro – Mammalian Cell Gene Mutation Test (OECD TG 476)

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1985/07/05-1985/07/15

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable well-documented study report which meets basic scientific principles: GLP.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Target gene:

Not applicable

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

S9 liver fractions from Aroclor exposed rats

Test concentrations with justification for top dose:

Tests (done in triplicate) with and without Metabolic Activation: Acetone (vehicle control), 0, 50, 150, 500, 1500, 5000 ug/plate
Vehicle control: 0.1 ml/plate acetone
Positive controls: 0.5ug/plate 2AA, 5ug/plate MNNG, 80ug/plate 9AA, 2ug/plate 2NF

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

0.1 ml/plate Acetone

True negative controls:

yes

Remarks:

non treated

Positive controls:

yes

Positive control substance:

other: TA 1537 (+S9 2-aminoanthracene) TA 1537 (-S9 9-aminoacridine); TA 98 (-S9 2-nitrofluorene) (+S9 2-aminoanthracene); TA100 (-S9 MNNG) (+S9 2-aminoanthracene); TA1535 (-S9 MNNG) (+S9 2-aminoanthracene); TA138(-S9 2-Nitrofluorene) (+S9 2-aminoanthracene)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar
DURATION
- Exposure duration: 72 hours


NUMBER OF REPLICATIONS:
- triplicate

DETERMINATION OF CYTOTOXICITY
- Method: reduction in the number of revertants and/or clearing of the background lawn of bacterial growth

Evaluation criteria:

The mutagenicity study is considered valid if the mean colony counts of the control values of the strains are within acceptable ranges, if the positive controls meet the criteria for a positive response and if no more than 5% of the plates are lost through contamination or other unforeseen events.

A test substance is considered to be positive in the bacterial gene mutation test if the mean number of revertant colonies on the test plates increase in a concentration-related manner and/or if it is a reproducible two-fold or more increase is observed compared to that on the negative control plates.

A test substance is considered negative in the bacterial gene mutation test if it produces neither a dose-related increase in the mean number of revertant colonies nor a reproducible positive response at any of the test points.

Positive results from the bacterial reverse mutation test indicate that a substance induces point mutations by base substitution for frameshifts in the genome of Salmonella typhimurium. Negative results indicate that under the test conditions, the test substance is not mutagenic.

Statistics:

The mean plate count and standard deviation for each dose point were determined. Any test value that was equal to or greater than two times the mean value of the concurrent vehicle control was considered to be a positive dose.

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

not cytotoxic up to 5,000ug/plate

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

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

not cytotoxic up to 5,000ug/plate

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

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

not cytotoxic up to 5,000ug/plate

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

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

not cytotoxic up to 5,000ug/plate

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

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

not cytotoxic up to 5,000ug/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Conclusions:

Interpretation of results: negative

Based on these data, classification under EU requirements for dangerous substances and preparations and under EU GHS guidelines is not required.

Executive summary:

Petrepar 120 was examined for mutagenic activity in the bacterial reverse mutation test using histidine-requiring Salmonella typhimurium strains TA 1535, 1537, 1538, 98, and 100 in the absence and presence of a liver S9 fraction for metabolic activation. The test was performed in triplicate using doses of 0, 50, 150, 500, 1500, 5000 ug/plate.  In all cases, Petrepar 120 did not induce any significant changes in the number of revertant colonies.  It is concluded in this study that Petrepar 120 is not a mutagenic agent and classification under EU requirements for dangerous substances and preparations guidelines is not required, nor is it required under EU GHS guidelines.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

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

Adequacy of study:

key study

Study period:

1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: “Acceptable, well-documented study report equivalent or similar to OECD guideline 473: GLP

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:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Target gene:

N/A

Species / strain / cell type:

primary culture, other: human lymphocytes from two male and one female donor

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

Experiment 1 without S9 (40.36, 57.66, 82.34 ug/ml);
Experiment 1 with S9 for 3 hours followed by 17 hour recovery (490, 700, 1000 ug/ml)
Experiment 2 without S9 20h treatment 0h recovery (22.52, 28.15, 35.18 ug/ml)
Experiment 2 with S9 for 3 hours followed by 17 hours recovery (640, 800, 1000 ug/ml)
Experiment 2 with S9 for 3 hours followed by 41 hours recovery (1000 ug/ml)
Experiment 3 without S9 for 20 hours treatment and 0 hours recovery (28.15, 35.19, 43.99 ug/ml)
Experiment 3 without S9 for 44 hours and 0 hours recovery (43.99 ug/ml)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

other: 4-nitroquinoline, cyclophosphamide

Evaluation criteria:

1) a statistically significant increase in the proportion of cells with structural aberrations (excluding gaps) occurred at one or more concentrations, and 2) the proportion of cells with structural aberrations at such doses exceeded normal range, and 3) the results confirmed in the second experiment. A positive result only at delayed harvest in Experiment 2 was to be taken as evidence of clastogenicity provided criteria 1 and 2 were met. Increases in numbers of cells with gaps or increases in the proportions of cells with structural aberrations, not exceeding the normal range or occurring only at very high or very toxic concentrations, were likely to be concluded as equivocal. Full assessment of the biological importance of such increases is likely to be possible with reference to data from other test systems. Cells with exchange aberrations or cells with greater than one aberration were to be considered of particular biological significance.

Key result

Species / strain:

primary culture, other: human peripheral blood lymphocytes

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Conclusions:

Interpretation of results: negative

The mammalian chromosomal aberration test to assess the genotoxicity of SHELLSOL D70 was negative. This finding does not warrant the classification of this test material as a genotoxin under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP) or under Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.

Executive summary:

The potential of SHELLSOL D70 to cause chromosome aberration was investigated in cultured human lymphocytes with and without the metabolic activation S9 system. Negative and positive control substances were include in both experiments to confirm the activity and sensitivity of the test systems.  In the first experiment, the maximum dose levels selected for chromosome analysis were 82.34 ug/ml and 1000 ug/ml, in the absence and presence of S9 respectively.  These dose levels caused inhibitions of the mitotic index of 57% and 30% respectively.  In the second experiment, the highest concentration used for chromosome analysis were 35,18 ug/ml and 1000 ug/ml in the absence and presence of S9 respectively, these gave a reduction in the mitotic index of 52% and 12% respectively.  In both Experiments 1 and 2 in the presence of S9; and in Experiment 2 in the absence of S9 only there were no significant increases in the frequency of the cells with structural aberrations in cultures treated with SHELLSOL D70.  Following treatment in Experiment 2 in the absence of S9 there was a significant increase in the frequency of structural aberrations at the lowest dose analyzed (22.52ug/ml).  Additional doses from Experiment 1 were analyzed (19.79 and 28.25 ug/ml) to confirm whether this effect was only apparent at low concentrations.  No increase in the frequency of structural aberrations was apparent at these concentrations.  In order to further clarify the findings seen in the initial experiments, a third experiment was performed in which there were no significant increases in the frequency of cells with structural aberrations in all cultures treated with SHELLSOL D70.  Since the increase in structural aberrations seen at 22.52 ug/ml in Experiment 2 was not apparent in other experiments at similar or higher concentrations, the effect was considered to be non-reproducible and of no biological importance.  Based on these results, it is concluded that SHELLSOL D70 did not induce chromosome aberrations in cultured lymphocytes when tested to its limit of toxicity in both the absence and presence of S9.  This finding does not warrant the classification of this test material as a genotoxin under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP) or under Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

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

Adequacy of study:

key study

Study period:

1982

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Well conducted study according to sound 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:

equivalent or similar to guideline

Guideline:

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

Deviations:

no

GLP compliance:

no

Type of assay:

mammalian cell gene mutation assay

Target gene:

TK+/ phenotype

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

TK+/ phenotype of L5178Y mouse lymphoma cells from subline 3.7.2C

Metabolic activation:

with and without

Metabolic activation system:

Aroclor

Test concentrations with justification for top dose:

up to was 1000 ug/mL in dimethylsulfoxide (maximum dose)

Vehicle / solvent:

dimethylsulfoxide

Untreated negative controls:

yes

Negative solvent / vehicle controls:

no

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

not specified

Details on test system and experimental conditions:

This assay was performed with the TK+/ phenotype of L5178Y mouse lymphoma cells from subline 3.7.2C using a minimum of eight test compound doses with and without metabolic activation by an Aroclor induced rat liver microsomal fraction. Appropriate negative, solvent, and positive controls were included with each assay. The test compound dose levels were determined by a preliminary multidose ranging study with the highest dose targeted to give approximately fifty to ninety percent inhibition of suspension cell growth depending on the solubility of the compound. C10-C13 isoalkanes achieved a homogeneous mixture at approximately 100 mg/ml in dimethylsulfoxide. The maximum dose selected for the mutagenicity test was 1000 ug/ml because it represents the limits of solubility of the test material.

Key result

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Exposure to eight graded doses of the test material in the presence of and in the absence of metabolic activation did not increase the induction of forward mutations in L5178Y mouse lymphoma cells at the T/K locus. Therefore C10-C13 isoalkanes are not considered to be mutagenic in this test system.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Conclusions:

Interpretation of results: negative with and without metabolic activation

Exposure to eight graded doses of the test material in the presence of and in the absence of metabolic activation did not increase the induction of forward mutations in L5178Y mouse lymphoma cells at the T/K locus. Therefore C10-C13 isoalkanes are not considered to be mutagenic in this test system.

Executive summary:

Exposure to eight graded doses of the test material in the presence of and in the absence of metabolic activation did not increase the induction of forward mutations in L5178Y mouse lymphoma cells at the T/K locus. Therefore C10-C13 isoalkanes are not considered to be mutagenic in this test system.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

In vivo genetic toxicity data is available for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics. An additional in vivo genetic toxicity test is available from structural analogue Hydrocarbons, C10-C12, isoalkanes, <2% aromatics. This data is read across to Hydrocarbons, C10 -C13, n-alkanes, <2% aromatics 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 genetic toxicity test listed below had a negative result for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics.

 

Genetic Toxicity in vivo – Mammalian Erythrocyte Micronucleus Test (equivalent/similar to OECD 474)

 

The read across genetic toxicity test listed below had a negative result for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics.

Genetic Toxicity in vivo – mammalian cell study: DNA damage and/or repair

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1990/10/24 - 1990/11/30

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: According to or similar to OECD Guideline 474. GLP

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 474 (Mammalian Erythrocyte Micronucleus Test)

GLP compliance:

yes

Type of assay:

micronucleus assay

Species:

mouse

Strain:

CD-1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
Source: Charles River Breeding Laboratories, Inc.
Sex: Male (65), Female (65)
Age at study initiation: Approximately 9-10 weeks
Weight at study initiation: 23-39g
Housing: Individually
Diet (e.g. ad libitum): Purina Certified Rodent 5002 chow (pellets), ad libitum
Water (e.g. ad libitum): Automatic watering system, ad libitum
Acclimation period: 7d

ENVIRONMENTAL CONDITIONS
Temperature (°F): 68-76
Humidity (%): 40-70%
Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

Corn oil was used. Dose volume did not exceed 1.0 ml/100 g bw.

Details on exposure:

The test material and the carrier were administered by oral gavage as a single dose. The carrier was dosed at a volume equal to the test material dose volume. The individual animal dose volumes did not exceed 1.0 ml/100 g body weight; animals were administered 1.0, 2.5, 5.0 g test material/ kg body weight. The positive control, cyclophosphamide was administered as a single dose of 40 mg/kg using water as a carrier.

Duration of treatment / exposure:

Animals were sacrificed 24, 48, and 72 hours after dose administration.

Frequency of treatment:

One dose was given at either 1.0, 2.5, 5.0 g test material/ kg body weight. Cyclophosphamide was dosed at 40 mg/kg.

Post exposure period:

Animals were sacrificed 24, 48, and 72 hours after dose administration.

No. of animals per sex per dose:

Male (65), Female (65) ; 5 Males and 5 Females per treatment group

Positive control(s):

The positive control, cyclophosphamide was administered as a single intraperitoneal injection (40 mg/kg) using water as a carrier.

Tissues and cell types examined:

Erythrocytes derived from femur bone marrow.

Details of tissue and slide preparation:

Immediately following the sacrifice of the animals, both femurs were removed and the bone marrow was removed and suspended in fetal bovine serum. After the suspension was centrifuged the pellet was resuspended and smears were prepared (two slides per animal).

Evaluation criteria:

Slides were stained using acridine orange; polychromatic erythrocytes (PCE) stained red/orange, nonchromatic erythrocytes (NCE) are unstained (dull green), and micronuclei stain bright yellow. Additional criteria for scoring micronuclei are a circular appearance and a diameter between 1/20 and 1/5 of the cell’s diameter. 1000 PCE from each animal were examined for the presence of micronuclei and the ratio of PCE to NCE was determined for each animal by counting 1000 erythrocytes (PCE and NCE).

Statistics:

Calculation of means and standard deviations of the micronuclei data and a test of equality of group means by a standard one way analysis of variance at each time period (ANOVA). When ANOVA was significant, comparisons of carrier control to dosed group means were made according to Duncan’s Multiple Range Test.

A standard regression analysis was performed to test for a dose response.
Residuals from the ANOVA were analyzed for normality by Wilk’s Criterion. The residuals were normally distributed (values were greater than 0.01 level of significance). Therefore nonparametric analysis was not performed.

Sexes were analyzed separately.

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

The positive control (cyclophosphamide) induced a statistically significant increase in the mean number of micronucleated polychromatic erythrocytes, indicating that the positive control was clastogenic and was responding in an appropriate manner. Carrier control values for the mean percent of polychromatic erythrocytes and for the mean number of micronucleated polychromatic erythrocytes are within the normal range for the corn oil control. MRD-90-874 did not induce a statistically significant decrease in the mean percent of polychromatic erythrocytes which is a measure of bone marrow toxicity. MRD-90-874 did not induce a statistically significant increase in the mean number of micronucleated polychromatic erythrocytes. MRD-90-874 did not induce a significant increase in the mean number of micronucleated polychromatic erythrocytes. MRD-90-874 was not cytotoxic at doses up to 5.0 g/kg to mouse bone marrow under the conditions of this test.

Conclusions:

Interpretation of results: negative
These data indicate that MRD-90-874 is not cytotoxic and is not clastogenic in CD-1 mouse bone marrow cells at doses up to and including 5.0 g/kg of body weight.

Executive summary:

The test material, MRD-90-874 was tested in the mammalian bone marrow micronucleus assay using CD-1 mice.  MRD-90-874 was tested at 24, 48, and 72 hour intervals following exposure and did not induce a statistically significant decrease in the mean percent of polychromatic erythrocytes or an increase in the mean number of micronucleated polychromatic erythrocytes.  Both the positive (cyclophosphamide) and the negative (carrier) controls behaved in an appropriate manner.  These data indicate that MRD-90-874 is not cytotoxic and is not clastogenic in CD-1 mouse bone marrow cells at doses up to and including 5.0 g/kg.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

In vitro and in vivo genetic toxicity data is available for Hydrocarbons, C10-C13, n-alkanes, <2% aromatics. Additional in vitro and in vivo data is available for structural analogues Hydrocarbons, C10-C12, isoalkanes, <2% aromatics, Hydrocarbons, C10-C13, isoalkanes, <2% aromatics and Hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, <2% aromatics. This data is read across to Hydrocarbons, C10 -C13, n-alkanes, <2% aromatics 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 study in bacteria

In a key study (Cepsa Quimica, 1985) equivalent/similar to OECD Guideline 471, the test material (Hydrocarbons, C10-C13, n-alkanes, <2% aromatics) was examined for mutagenic activity in the bacterial reverse mutation test using histidine-requiring Salmonella typhimurium strains TA 1535, 1537, 1538, 98, and 100 in the absence and presence of a liver S9 fraction for metabolic activation. The test was performed in triplicate using doses of 0, 50, 150, 500, 1500, 5000 ug/plate.  In all cases, the test material did not induce any significant changes in the number of revertant colonies.  It is concluded in this study that the test material is not a mutagenic agent and classification under EU requirements for dangerous substances and preparations guidelines is not required, nor is it required under EU GHS guidelines.

 

In vitro Chromosome Aberration in Mammalian Cells

In a key OECD Guideline 473 study (Shell, 1998), the potential of the test material (Hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, <2% aromatics) to cause chromosome aberration was investigated in cultured human lymphocytes with and without the metabolic activation S9 system. Negative and positive control substances were include in both experiments to confirm the activity and sensitivity of the test systems.  In the first experiment, the maximum dose levels selected for chromosome analysis were 82.34 ug/ml and 1000 ug/ml, in the absence and presence of S9 respectively.  These dose levels caused inhibitions of the mitotic index of 57% and 30% respectively.  In the second experiment, the highest concentration used for chromosome analysis were 35,18 ug/ml and 1000 ug/ml in the absence and presence of S9 respectively, these gave a reduction in the mitotic index of 52% and 12% respectively.  In both Experiments 1 and 2 in the presence of S9; and in Experiment 2 in the absence of S9 only there were no significant increases in the frequency of the cells with structural aberrations in cultures treated with the test material.  Following treatment in Experiment 2 in the absence of S9 there was a significant increase in the frequency of structural aberrations at the lowest dose analyzed (22.52ug/ml).  Additional doses from Experiment 1 were analyzed (19.79 and 28.25 ug/ml) to confirm whether this effect was only apparent at low concentrations.  No increase in the frequency of structural aberrations was apparent at these concentrations.  In order to further clarify the findings seen in the initial experiments, a third experiment was performed in which there were no significant increases in the frequency of cells with structural aberrations in all cultures treated with the test material.  Since the increase in structural aberrations seen at 22.52 ug/ml in Experiment 2 was not apparent in other experiments at similar or higher concentrations, the effect was considered to be non-reproducible and of no biological importance.  Based on these results, it is concluded that Hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, <2% aromatics did not induce chromosome aberrations in cultured lymphocytes when tested to its limit of toxicity in both the absence and presence of S9.

 

In vitro Gene Mutation study in Mammalian Cells

In a key study (Chevron, 1982), exposure to eight graded doses of the test material (Hydrocarbons, C10-C13, isoalkanes) in the presence of and in the absence of metabolic activation did not increase the induction of forward mutations in L5178Y mouse lymphoma cells at the T/K locus. Therefore C10-C13 isoalkanes are not considered to be mutagenic in this test system.

   

In Vivo

 

In a key study (Exxon, 1991) equivalent/similar to OECD Guideline 474, the test material (Hydrocarbons, C10-C13, n-alkanes, <2% aromatics) was tested in the mammalian bone marrow micronucleus assay using CD-1 mice.  The test material was tested at 24, 48, and 72 hour intervals following exposure and did not induce a statistically significant decrease in the mean percent of polychromatic erythrocytes or an increase in the mean number of micronucleated polychromatic erythrocytes.  Both the positive (cyclophosphamide) and the negative (carrier) controls behaved in an appropriate manner.  These data indicate that Hydrocarbons, C10-C13, n-alkanes, <2% aromatics are not cytotoxic and are not clastogenic in CD-1 mouse bone marrow cells at doses up to and including 5.0 g/kg.

In a supporting study (Shell, 1998), C3H mice were treated cutaneously with a range of doses (1x25ml to 3x25ml per animal; ca. 900-2750mg/kg) of either n-decane or n-dodecane for periods ranging from 24 to 72 hours.  The DNA was isolated for the epidermis of the treated skin and subjected to 32P-postlabeling analysis.  A group of untreated control animals was untreated and maintained under the same conditions as the exposed groups.  For a positive control, a group of mice was treated with benzo(a)pyrene in 25ul dodecane at a level of 1ug per animal, a dose which is about 2 orders of magnitude lower than a single carcinogenic dose of benzo(a)pyrene for this species.  For a positive control test of the 32P-postlabelling assay, DNA from mice treated with benzo(a)pyrene in THF was used.  32P-Postlabelling analysis of the epidermal DNA from mice treated with either n-decane or n-dodecane at each of the exposure levels and durations showed an absence of radioactive spots or diagonal radioactive zones which could have corresponded to adducts arising from decane.  The positive control adduct was easily detectable.  The limit of detection of the procedure was about 1 adduct in 109 nucleotides using 10mg samples of DNA.  The absences of adducts with this level of sensitivity of adduct detection indicates that n-decane and n-dodecane do not possess genotoxic activity in vivo.

Justification for classification or non-classification

The negative results using in vitro and in vivo genotoxicity assays from Hydrocarbons, C10-C13, n-alkanes, <2% aromatics and structural analogues do not warrant the classification of Hydrocarbons, C10-C13, n-alkanes, <2% aromatics as genotoxic under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP).