Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Gene mutation (Bacterial reverse mutation assay / Ames test): negative with and without activation in S. typhimurium strains TA 98, TA100, TA1535, TA1537 and TA 1538 (similar to OECD TG 471) (Safepharm Laboratories, 1996)

Gene mutation (Bacterial reverse mutation assay / Ames test): the related substance Fatty alcohol blend (containing 40.77% C8 and 55.3% C10) was negative with and without activation in S. typhimurium strains TA 98, TA100, TA1535, TA1537 and TA 1538 (similar to OECD TG 471) (Iglesias, 2002b)

Cytogenicity in mammalian cells: the related substance C12 and 13 alcohols; linear and monobranched, type 2: negative in CHO cells (OECD TG 473) (Sasol, 1998)
Cytogenicity in mammalian cells: the related substance Docosan-1-ol was negative with and without activation in Chinese hamster ovary cells (similar to OECD TG 473) (Iglesias, 2002b).
Mutagenicity in mammalian cells: the related substance Docosan-1-ol was negative with and without activation in Chinese hamster lung V79 cells (similar to OECD TG 476) (Iglesias, 2002b).

Mutagenicity in mammalian cells: the related substance Fatty alcohol blend (containing 40.77% C8 and 55.3% C10): negative with and without activation in L5178Y mouse lymphoma cells (similar to OECD TG 476) (Inveresk, 1992).

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1998
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
GLP compliance:
yes (incl. certificate)
Type of assay:
in vitro mammalian chromosome aberration test
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Metabolic activation system:
Aroclor induced rat liver S9
Test concentrations with justification for top dose:
Test 1: 0.1 - 500 µg/ml; Test 2: with S9 1 -50 µg/ml, without S9 0.5 - 20 µg/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: none given in report. Standard solvent
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
without activation
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with activation
Details on test system and experimental conditions:
ACTIVATION: 1 ml Aroclor induced rat liver S9 mix, NADP as cofactor
METHOD OF APPLICATION: in medium

DURATION

Exposure duration: Test 1: +S9 3 hours, -S9 18 hours, Test 2: +S9 3 hours; -S9  18 or 32 hours 
- Expression time (cells in growth medium): 18 hours
- Fixation time (start of exposure up to fixation or harvest of cells): Test 1 18 hours; Test 2 18 or 32 hours

SPINDLE INHIBITOR (cytogenetic assays): Colcemid (final concentration 0.1 mM)

TAIN (for cytogenetic assays): Giesma

NUMBER OF REPLICATIONS: Duplicate cultures, independent repeat assay

NUMBER OF CELLS EVALUATED: 100 per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; other: observation of culture

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes
- Other: interstitial deletions
Evaluation criteria:
CRITERIA FOR EVALUATING RESULTS: The test is considered positive if the aberration frequency of at least one concentration is significantly above concurrent control frequencies.
Statistics:
Fisher's exact probability test (two-sided)
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
+S9 40 µg/ml; -S9 15 µg/ml
Vehicle controls validity:
valid
Untreated negative controls validity:
not valid
Positive controls validity:
valid
Additional information on results:
GENOTOXIC EFFECTS: - With and without metabolic activation:  
There were  no statistically significant increase in total numbers of chromosome aberrations at any dose level tested
There was no increase in the incidence of  polyploids or endoreduplicates.
PRECIPITATION CONCENTRATION: 125 µg/ml. 
MITOTIC INDEX: The mitotic index was measured on 1000 cells and was always >40% of control levels and usually >50% for the dose levels which   were scored for chromosome aberrations. 
STATISTICAL RESULTS: Fischers exact probability test (two-sided) did not indicate any significant difference between test and control groups.
Remarks on result:
other: No mutagenic potential

Chromosome aberration assay: Test 1

Treatment time 18 hrs

Treatment

Activation

Concentration µg/ml

Number of cells

Total - gaps

Total + gaps

Negative control DMSO

-MA

0

200

0

4

Test substance

 

 

-MA

2.5

200

0

6

-MA

5

200

0

6

-MA

10

200

2

8

Positive control Mitomycin C

-MA

0.025

200

48

48

Treatment time 3 hrs

Treatment

Concentration µg /ml

Number of cells

Total - gaps

Total + gaps

Negative control DMSO

+MA

0

200

0

2

Test substance

 

 

+MA

10

200

1

6

+MA

20

200

1

5

+MA

30

200

1

5

Positive control Cyclophosphamide

+MA

3.75

200

48

48

Chromosome aberration assay: Test 2

Treatment

-MA

Concentration 

µg /ml

Number of cells

Total - gaps

Total + gaps

Treatment time 18 hrs

Negative control DMSO

-MA

0

200

1

5

Test substance

 

 

-MA

5

200

2

8

-MA

7.5

200

0

5

-MA

10

200

1

4

Positive control Mitomycin C

-MA

0.025

200

54

54

Treatment time 3 hrs, incubation 18 hours

Treatment

Concentration

µg /ml

Number of cells

Total - gaps

Total + gaps

Negative control DMSO

+MA

0

200

0

8

Test substance

 

 

+MA

10

200

1

2

+MA

20

200

1

7

+MA

30

200

1

7

Positive control Cyclophosphamide

+MA

3.75

200

105

105

Treatment time 32 hrs

Treatment

Concentration

µg /ml

Number of cells

Total - gaps

Total + gaps

Negative control DMSO

-MA

0

200

0

8

Test substance

-MA

10

200

0

7

Treatment time 3 hrs, incubation 32 hours

Treatment

Concentration 

µg /ml

Number of cells

Total - gaps

Total + gaps

Negative control DMSO

+MA

0

200

2

3

Test substance

+MA

30

200

0

5

 

Conclusions:
C12 AND 13 ALCOHOLS; LINEAR AND MONOBRANCHED, TYPE 2 (also known as Compound 33A) has been tested in a valid study according to OECD TG 473 and under GLP in CHO K1 cells. The test substance did not increase the incidence of chromosome aberrations in Chinese hamster ovary cells at dose levels up to cytotoxic concentrations in the presence or absence of metabolic activation. The result of the repeat experiment confirmed that of the initial assay. It is concluded that the test substance is negative for the induction of chromosome aberrations under the conditions of this test.
Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
19-Sep-1996 to 24-Sep-1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
The restriction was that no cross linking strain was used, so does not comply with current guidelines.
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
yes
Remarks:
(no TA102 or E coli WP2 uvrA, 2-AA only positive control with S9)
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay
Target gene:
histidine
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:
Aroclor 1254-induced rat liver S9
Test concentrations with justification for top dose:
Test 1: 15(-S9 only), 50, 150, 500, 1500, and 5000 µg/plate; Test 2: 50, 150, 500, 1500, and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Well-known solvent
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
TA100 (3 ug/plate) and TA1535 (5 ug/plate) without S9
Positive control substance:
N-ethyl-N-nitro-N-nitrosoguanidine
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
TA1537 (8 ug/plate) without S9
Positive control substance:
9-aminoacridine
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
TA1538 (5 ug/plate) without S9
Positive control substance:
other: 4-nitro-o-phenylenediamine
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
TA98 (0.2 ug/plate) without S9
Positive control substance:
4-nitroquinoline-N-oxide
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
all strains (0.5, 1 or 2 ug/plate) with S9
Positive control substance:
other: 2-aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium; in agar (plate incorporation)

DURATION
- Exposure duration: approximately 48 hours

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Thinning or absence of background lawn of non-revertant cells

OTHER: The mutation experiment was repeated on a separate day using fresh cultures and fresh test material formulations.
Evaluation criteria:
After incubation, numbers of revertant colonies were counted. For a substance to be considered positive in this test system, it should have induced a dose-related and statistically significant increase in mutation rate in one or more strains of bacteria in the presence and/or absence of S9 in both experiments at sub-toxic dose levels. For a negative result the numbers of induced revertants should be less than two fold compared to controls.
Statistics:
Dunnetts test was used and showed no statistically significant differences between test and control plates.
Key result
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and 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
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
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
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no data
- Effects of osmolality: no data
- Evaporation from medium: no data
- Water solubility: no data
- Precipitation: precipitation of test substance was observed at dose levels of 1500 ug/plate and above. Plates were counted manually at 1500 ug/plate and above.
- Other confounding effects: no data

RANGE-FINDING/SCREENING STUDIES: Strain TA 100 was exposed to the test substance at concentrations of 0, 50, 150, 500, 1500 and 5000 ug/plate in a preliminary toxicity study.

COMPARISON WITH HISTORICAL CONTROL DATA: no data

CYTOTOXIC CONCENTRATION: Slight cytotoxicity was indicated in a preliminary toxicity screen with TA100 at dose levels >= 500 ug/plate without 
metabolic activation. In the actual mutation study there was no evidence of cytotoxicity up to 5000 ug/plate with or without S9.
Remarks on result:
other: No mutagenic potential

Table 1:Number of revertants per plate (mean of 3 plates) for Test 1

 

Conc.
[
µg /plate]

 

[TA 100]

[TA 1535]

[TA 1538]

[TA 98]

[TA 1537]

-MA

+MA

Cytotoxic
(yes/no)

- MA

+MA

Cytotoxic
(yes/no)

- MA

+MA

Cytotoxic
(yes/no)

-MA

+ MA

Cytotoxic
(yes/no)

-MA

+ MA

Cytotoxic
(yes/no)

0*

72

116

no

14

15

no

14

25

no

20

29

no

6

9

no

15

71

-

no

12

-

no

11

-

no

18

-

no

7

-

no

50

65

112

no

12

15

no

11

27

no

17

24

no

6

12

no

150

64

98

no

12

12

no

12

22

no

18

35

no

7

12

no

500

57

89

no

12

14

no

11

28

no

21

27

no

8

10

no

1500

55

72

no

12

17

no

11

22

no

19

27

no

7

9

no

5000

52

74

no

12

11

no

12

22

no

23

30

no

10

7

no

Positive control

597

831

no

348

195

no

636

488

no

217

485

no

550

282

no

*solvent control with DMSO

 

Table 2: Number of revertants per plate (mean of 3 plates) for Test 2

 

Conc.
[
µg /plate]

 

[TA 100]

[TA 1535]

[TA 1538]

[TA 98]

[TA 1537]

-MA

+MA

Cytotoxic
(yes/no)

-MA

+MA

Cytotoxic
(yes/no)

-MA

+MA

Cytotoxic
(yes/no)

-MA

+MA

Cytotoxic
(yes/no)

-MA

+MA

Cytotoxic
(yes/no)

0*

86

85

no

19

12

no

11

16

no

15

24

no

7

9

no

50

67

83

no

15

13

no

10

14

no

19

28

no

7

8

no

150

70

66

no

11

9

no

9

13

no

20

29

no

5

7

no

500

66

71

no

17

10

no

9

14

no

15

26

no

6

11

no

1500

66

72

no

9

11

no

7

12

no

13

20

no

5

8

no

5000

66

58

no

9

10

no

6

13

no

12

23

no

7

9

no

Positive control

459

1005

no

211

208

no

524

344

no

175

345

no

762

266

no

*solvent control with DMSO

 

 



Conclusions:
In a reliable study, conducted in accordance with OECD guideline 471 and under GLP, the C14 alcohol Kalcol 4098 did not increase the reverse mutation rate in histidine dependent bacterial strains of Salmonella typhimurium in the presence or absence of metabolic activation at concentrations up to 5000 µg/plate. The top concentration was not cytotoxic. It is concluded that the test substance is not mutagenic to bacteria under the conditions of the test.
Executive summary:

In the bacterial mutagenicity study, the ability of tetradecan-1 -ol to induce mutations in bacteria was tested.

In test I, 50, 150, 500, 1500, and 5000 µg/plate of test material were applied to S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 1538 without metabolic activation. In test II, the same concentrations of the test material were applied to

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 1538, in the presence and absence of metabolic activation system. The exposure duration was approximately 48 hours.

After incubation, numbers of revertant colonies were counted. For a substance to be considered positive in this test system, it should have induced a dose-related and statistically significant increase in mutation rate in one or more strains of bacteria in the presence and/or absence of metabolic activation in both experiments at sub-toxic dose levels. For a negative result the numbers of induced revertants should be less than two fold compared to controls.

The study reports tetradecan-1 -ol to be not mutagenic to S. typhimurium when tested up to limit concentration. The study was conducted according to an appropriate OECD test guideline, with acceptable restriction. The restriction was that no cross linking strain was used, so does not comply with current guidelines. It was compliant with GLP.

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
no cross linking strain was used
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
yes
Remarks:
(no TA102 or E. coli WP2 uvrA; 2-aminoanthracene only positive control with metabolic activation)
Principles of method if other than guideline:
Well-conducted study according to a protocol very similar to OECD guideline 471
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Target gene:
histidine
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:
liver microsomal fractions from male rats prepared by "established methods"
Test concentrations with justification for top dose:
10, 100, 333, 667, and 1000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: solubility
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA100, TA1535 without metabolic activation
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 4-nitro-O-phenylenediamine
Remarks:
TA98, TA1537, TA1538 without metabolic activation
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: aminoanthracene
Remarks:
all strains with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: none
- Exposure duration: no data

NUMBER OF REPLICATES:
- two independent experiments, both with and without metabolic activation
- each concentration (including controls) tested in triplicate

DETERMINATION OF CYTOTOXICITY
- Method: no data
Evaluation criteria:
To be considered positive in TA100, >=2x increase in revertants over spontaneous rate; in TA98, TA1535, TA1537 and TA1538, >=3x increase; alternatively a concentration-dependent increase irrespective of 2- or 3-fold increase
Statistics:
none
Key result
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: not determined, but number of revertants reduced in TA 98 at 1000 µg/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:
other: not determined, but number of revertants not reduced at 1000 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no data
- Effects of osmolality: no data
- Evaporation from medium: no data
- Water solubility: insoluble
- Precipitation: no data
- Other confounding effects: no data

RANGE-FINDING/SCREENING STUDIES: yes, no data presented

COMPARISON WITH HISTORICAL CONTROL DATA: no

ADDITIONAL INFORMATION ON CYTOTOXICITY: none
Remarks on result:
other: No mutagenic potential

Table 1 Revertants per plate (mean of 3 plates)

Concentration µg/plate

TA 98

TA100

TA1535

TA1537

TA1538

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

Negative control

15.3

17.0

82.3

833.7

5.7

8.0

5.0

4.0

14.3

16.7

0*

15.3

15.7

83.3

77.3

8.3

8.3

7.0

5.0

14.7

15.0

10.0

14.0

19.0

70.7

80.3

10.7

9.7

3.0

5.0

14.3

14.0

100.0

9.3

15.3

85.0

82.0

7.7

9.0

4.0

4.3

14.7

15.7

333.3

11.7

15.7

80.0

79.7

8.0

12.3

4.7

5.0

15.0

15.3

666.6

12.0

12.7

74.0

82.3

8.3

6.3

4.3

5.3

14.0

15.3

1000

6.7

8.0

76.0

86.3

4.7

3.3

3.7

5.0

13.7

14.7

Positive control

1573

2337.7

1158

2414

601.7

345

109.7

85.3

1980

495.7

* Solvent control with

Conclusions:
In a valid and reliable study, behenyl alcohol (C22) did not increase the reverse mutation rate in histidine dependent bacterial strains of Salmonella typhimurium in the presence or absence of metabolic activation at concentrations up to and including 1000 µg/plate. It is concluded that the test substance is negative for mutagenicity in bacteria under the conditions of the test.
Executive summary:

In the bacterial mutagenicity study, S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 1538 bacteria were incubated with 10, 100, 333, 667, and 1000 µg/plate of test material, with and without metabolic activation. The test material did not increase the reverse mutation rate in histidine dependent bacterial strains of Salmonella typhimurium in the presence or absence of metabolic activation up to limit concentration. The study concludes that the test substance is negative for mutagenicity in bacteria under the conditions of the test. The study was conducted according to a test protocol that is comparable to the appropriate OECD test guideline, with the exception that no cross linking strain was used. Read across to the registered substance is considered scientifically justified.

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not stated
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
without detailed documentation
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
no
Principles of method if other than guideline:
Well-conducted study according to protocol very similar to OECD guideline 473
GLP compliance:
not specified
Type of assay:
in vitro mammalian chromosome aberration test
Target gene:
not applicable
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
- Type and identity of media: minimum essential medium
- Properly maintained: no data
- Periodically checked for Mycoplasma contamination: no data
- Periodically checked for karyotype stability: no data
- Periodically "cleansed" against high spontaneous background: no data
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
liver microsomal fraction from male rats prepared according to Ames et al., 1977
Test concentrations with justification for top dose:
0.6, 10.0 and 20.0 µg/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: solubility
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
without metabolic activation
Positive control substance:
ethylmethanesulphonate
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
with metabolic activation
Positive control substance:
cyclophosphamide
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: not applicable
- Exposure duration: 4 hours
- Expression time (cells in growth medium): not applicable
- Selection time (if incubation with a selection agent): not applicable
- Fixation time (start of exposure up to fixation or harvest of cells): 7 and 24 (or 28) hours at 20 µg/ml, 18 hours at 0.6, 10 and 20 µg/ml

SPINDLE INHIBITOR (cytogenetic assays): Colcemid, 0.2 µg/ml

STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS: 2 cultures per concentration

NUMBER OF CELLS EVALUATED: 100 per slide, 200 per concentration

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index
Evaluation criteria:
To be considered positive, either a statistically significant, concentration-related increase in the number of structural chromosome aberrations, or a statistically significant positive response at one of the concentrations
Statistics:
Chi-squared test performed for cells with aberration (excluding gaps)
Key result
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: presumably >20 µg/ml
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no data
- Effects of osmolality: no data
- Evaporation from medium: no data
- Water solubility: insoluble
- Precipitation: no data
- Other confounding effects: no data

RANGE-FINDING/SCREENING STUDIES: yes, but no data presented

COMPARISON WITH HISTORICAL CONTROL DATA: no data

ADDITIONAL INFORMATION ON CYTOTOXICITY: at 20 µg/ml, mitotic index not reduced, plating efficiency not reduced
Remarks on result:
other: No mutagenic potential

Table 1 Cytogenicity: 7 hour fixation. Aberrations in 200 cells

Activation

Concentration µg/ml

Percent aberrant cells

incl gaps

excl gaps

exchanges

Without

0*

4.0

1.5

0

20

2.5

0.5

0

With

0*

4.0

1.5

0

20

7.0

2.5

0

* Solvent control with ethanol

** Only 100 cells counted for positive controls

 

Table 2 Cytogenicity: 18 hour fixation. Aberrations in 200 cells

Activation

Concentration µg/ml

Percent aberrant cells

incl gaps

excl gaps

exchanges

Without

Negative control

5.5

1.5

0

0*

4.0

1.5

0.5

0.6

4.5

2.0

0

10

4.0

1.0

0.5

20

3.0

0.5

0

Positive control**

12.0

9.0

4.0

With

Negative control

2.5

1.5

0

0*

2.5

1.5

0.5

0.6

5.5

3.0

0.5

10

4.0

2.5

0

20

4.0

2.5

0.5

Positive control**

16.0

13.0

5.5

* Solvent control with ethanol

** Only 100 cells counted for positive controls

Table 3 Cytogenicity: 18 hour fixation. Aberrations in 200 cells

Activation

Concentration µg/ml

Percent aberrant cells

incl gaps

excl gaps

exchanges

Without

0*

6.0

2.5

0.5

20

3.5

2.0

0

With

0*

1.0

0.5

0

20

4.0

2.5

0.5

* Solvent control with ethanol

** Only 100 cells counted for positive controls

Conclusions:
In a reliable study, according to a protocol that is similar to OECD 473, behenyl alcohol (C22) did not increase the incidence of chromosome aberrations in Chinese hamster V79 cells in the presence or absence of metabolising fraction at concentrations up to 20 µg/ml. There was no evidence of cytotoxicity at this dose level.
Executive summary:

In an in vitro chromosome aberration study, Chinese hamster lung fibroblasts (V79) were incubated with 0.6, 10.0 and 20.0 µg/ml of test material dissolved in ethanol for 4 hours, with and without metabolic activation.

The test substance did not increase the incidence of chromosome aberrations in Chinese hamster V79 cells in the presence or absence of metabolic activation when tested up to limit concentration. There was no evidence of cytotoxicity at this dose level. The study was comparable to guideline without detailed documentation (publication). It is considered that read across to the registered substance is valid and scientifically justifiable.

Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay
Species / strain / cell type:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Metabolic activation system:
Aroclor induced rat liver S9
Test concentrations with justification for top dose:
Toxicity assay: 0.4, 4.3, 43.2, 432, 4320 µg/ml; Mutagenicity assay: 9.4, 18.8, 37.5, 75, 150, 300 µg/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO was used in toxicity assay, acetone in mutagenicity assay
- Justification for choice of solvent/vehicle: due to impurity peaks in the chromatograms, solvent was changed to acetone.
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
without metabolic activation
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
3-methylcholanthrene
Remarks:
with metabolic activation
Details on test system and experimental conditions:
ACTIVATION: 1.0 ml S9 mix containing 10% S9 and cofactors NADP and glucose-6-phosphate added to give final volume of 10 ml

METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: none

- Exposure duration: 4 hours

- Expression time (cells in growth medium): 2 days

- Selection time (if incubation with a selection agent): 11-14 days


SELECTION AGENT (mutation assays): trifluorothymidine

NUMBER OF REPLICATIONS: duplicate cultures, independent repeat experiment

DETERMINATION OF CYTOTOXICITY
- Method: other: cloning efficiency
Evaluation criteria:
A substance was considered positive if there was an increase of at least 1.7 fold in at least one of the highest doses was significant and associated with an increase in mutant numbers and/or an upward trend in the remaining doses, in two experiments under the same activation conditions.
Statistics:
Statistical evaluation was performed if marginal responses were recorded.
Key result
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
43.2 µg/ml
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid

Table 1 Experiment 1 Mutant frequency (average of 3 plates per culture)

Concentration µg/ml

Relative total growth

%

Mean mutant count

(MC)

Mutant fraction x 10¿¿

Increase over control

-MA

+MA

-MA

+MA

-MA

+MA

-MA

+MA

Solvent control

81

92

19

24

27

36

 

-

 

 

-

112

103

32

25

35

36

102

101

25

31

30

35

104

103

26

27

31

32

Positive control

75

34

182

151

264

299

8.6

7.6

63

38

150

135

268

235

9.4

129

85

17

30

21

36

1.0

0.9

100

88

29

18

39

26

18.8

111

92

27

21

31

25

1.2

0.7

109

71

39

19

44

24

37.5

72

92

18

24

25

34

0.9

1.0

79

71

27

27

33

35

75

-

-

NP

NP

-

-

-

-

-

-

NP

NP

-

-

150

-

-

NP

NP

-

-

-

-

-

-

NP

NP

-

-

300

-

-

NP

NP

-

-

-

-

-

-

NP

NP

-

 

NP = Not plated, too toxic for assessment

Table 2 Experiment 2 Mutant frequency (average of 3 plates per culture)

Concentration µg/ml

Relative total growth

%

Mean mutant count

(MC)

Mutant fraction x 10¿¿

Increase over control

-MA

+MA

-MA

+MA

-MA

+MA

-MA

+MA

Solvent control

94

99

25

35

28

43

-

-

113

110

30

41

26

43

92

99

29

31

28

37

-

93

-

30

-

37

Positive control

84

20

152

107

182

315

7.5

7.7

83

21

172

122

221

298

10

103

-

31

NP

33

-

1.3

-

98

-

32

NP

39

-

20

128

-

30

NP

29

-

1.1

-

86

-

28

NP

29

-

30

93

90

21

27

26

30

1.1

0.9

91

79

29

40

32

39

40

57

90

34

38

48

42

1.3

0.9

84

94

20

29

22

33

50

32

91

20

31

26

31

1.0

0.8

29

93

21

32

30

36

60

-

71

NP

36

-

36

-

0.8

-

56

NP

24

-

28

70

-

37

NP

22

-

25

-

0.6

-

23

NP

25

-

26

80

-

-

NP

NP

-

-

-

-

-

-

NP

NP

-

-

NP = Not plated: 3 highest dose levels, too toxic for assessment

Conclusions:
Fatty alcohol blend has been tested according to a protocol that is similar to OECD 476 and under GLP. No increase in the mutant frequency was observed with or without metabolic activation in either the initial or repeat experiment up to cytotoxic concentrations. Solvent and positive controls gave expected results. It is concluded that the test substance is negative for mutagenicity to bacteria under the conditions of the test.
Executive summary:

Fatty alcohol blend has been tested according to a protocol that is similar to OECD 476 and under GLP. No increase in the mutant frequency was observed with or without metabolic activation in either the initial or repeat experiment up to cytotoxic concentrations. Solvent and positive controls gave expected results. It is concluded that the test substance is negative for mutagenicity to bacteria under the conditions of the test.

The in vitro and in vivo data available for members of the category and supporting substances indicate that the C6-24 alcohols are not genotoxic. In addition, the category of LCAAs under consideration does not contain any structural elements that are of concern for potential mutagenic activity.

Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Principles of method if other than guideline:
Well-conducted study according to a protocol very similar to OECD guideline 476
GLP compliance:
not specified
Type of assay:
mammalian cell gene mutation assay
Target gene:
HGPRT
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
- Type and identity of media: no data
- Properly maintained: no data
- Periodically checked for Mycoplasma contamination: no data
- Periodically checked for karyotype stability: no data
- Periodically "cleansed" against high spontaneous background: no data
Metabolic activation:
with and without
Metabolic activation system:
no data, but for Ames test, liver microsomal fractions from male rats prepared by "established methods"
Test concentrations with justification for top dose:
2.0, 7.5, 15.0, and 20.0 ug/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: ethanol, final concentration in culture medium <=1% v/v
- Justification for choice of solvent/vehicle: solubility
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
without metabolic activation
Positive control substance:
ethylmethanesulphonate
Remarks:
1.0 ug/ml
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
with metabolic activation
Positive control substance:
7,12-dimethylbenzanthracene
Remarks:
15.4 ug/ml
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: not applicable
- Exposure duration: 4 hours
- Expression time (cells in growth medium): no data
- Selection time (if incubation with a selection agent): no data
- Fixation time (start of exposure up to fixation or harvest of cells): no data

SELECTION AGENT (mutation assays): thioguanine

NUMBER OF REPLICATIONS:
- 2 independent experiments, both with and without metabolic activation

NUMBER OF CELLS EVALUATED: no data

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth
Evaluation criteria:
To be considered positive, statistically significant concentration-related increase in mutant frequency, or a reproducible and statistically significant positive response for at least one concentration
Statistics:
no data
Key result
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: presumably >20 µg/ml
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no data
- Effects of osmolality: no data
- Evaporation from medium: no data
- Water solubility: insoluble
- Precipitation: no data
- Other confounding effects: no data

RANGE-FINDING/SCREENING STUDIES: yes, but no data presented

COMPARISON WITH HISTORICAL CONTROL DATA: no data

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- With metabolic activation: mean relative cell survival over the test concentrations ranged from 89.1% (20 ug/ml) to 93.8% (15 ug/ml).
- Without metabolic activation: mean relative cell survival ranged from 96% (15 ug/ml) to 120.2 % (20 ug/ml).
Remarks on result:
other: No mutagenic potential

Table 1 Results of mutagenicity in V79 cells (mean of 2 cultures)

Concentration µg/ml

Mean relative cell survival (%)

Mean mutants per culture

Mutant colonies per 10 E06 cells

-MA

+MA

-MA

+MA

-MA

+MA

Negative

105.1

98.2

2.7

4.8

8.65

47.4

0*

100

100

4.5

2.1

14.7

8.75

2

101.3

92.55

4.1

6.3

12.5

21.65

7.5

102.4

93.6

5.4

4.9

16.1

15.75

15

96.0

93.8

3.4

1.7

12.3

6.35

20

120.2

89.1

4.3

4.4

17.9

16.95

Positive control

67.3

104.6

156.7

39.1

1143.7

163.4

Conclusions:
In a reliable study, behenyl alcohol (C22) did not increase the gene mutation rate in Chinese hamster V79 cells in the presence or absence of metabolic activation at concentrations up to 20 ug/ml. It is concluded that the test substance is negative for mutagenicity in mammalian cells under the conditions of this test.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Mouse micronucleus study: the related substance dodecan-1-ol was negative in mice after oral administration (gavage) (OECD 474) (Henkel, 1992).
Mouse micronucleus study: the related substance docosan-1-ol was negative after oral administration (similar to OECD TG 474) (Iglesias, 2002b).

Micronucleus study in mice: the related substance Fatty alcohol blend (containing 40.77% C8 and 55.3% C10) was negative after oral administration (Inveresk, 1992).

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
key study
Study period:
26 November 1991 to 11 February 1332
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
only 1000 PCE per animal were scored for micronuclei
GLP compliance:
yes
Type of assay:
micronucleus assay
Species:
mouse
Strain:
CD-1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Limited, Manston Road, Kent

- Age at study initiation: 5-7 weeks

- Weight at study initiation: 27-30 g (males) 18-23 g (females)

- Assigned to test groups randomly: yes

- Fasting period before study: no information

- Housing: individually in polypropylene/stainless steel cages

- Diet: ad libitum

- Water: ad libitum

- Acclimation period: at least 10 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19

- Humidity (%): 38

- Air changes (per hr): no information

- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil

- Justification for choice of solvent/vehicle: none given; standard vehicle

- Concentration of test material in vehicle: sufficient to give required dose in appropriate volume of vehicle
- Amount of vehicle (if gavage or dermal): 10 mg/ml/day
Duration of treatment / exposure:
Animals were dosed on three consecutive days.
Frequency of treatment:
daily
Post exposure period:
96 hours
Dose / conc.:
500 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Dose / conc.:
2 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
5 (positive control, low and mid dose) or 10 (vehicle control, high dose)
Control animals:
yes, concurrent vehicle
Positive control(s):
- Positive control substance: cyclophosphamide

- Justification for choice of positive control(s): none given - standard positive control

- Route of administration: no information

- Doses / concentrations: 40 mg/ kg bw
Tissues and cell types examined:
Bone marrow
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: no deaths occurred in preliminary toxicity assay

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): Animals dosed at 0, 24 and 48 hours; samples taken at 72 and 96 hours

DETAILS OF SLIDE PREPARATION: Air dried slides were fixed in methanol then stained with 1% May-Grunwald for 5 minutes then counterstained in 15% Giesma for 15 minutes

METHOD OF ANALYSIS: 1000 PCE scored for micronuclei; PCE/NCE ratio was determined for 300 cells, using x 1000 oil immersion objective


Evaluation criteria:
An increase in micronucleus frequency of greater than 0.28%.
Statistics:
No statistical evaluation described.
Key result
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Negative controls validity:
not applicable
Positive controls validity:
valid

Table 1 Results of in vivo micronucleus study

 

Treatment

mg/kg.bw /day

 

Time of dosing (h)

 

Time of sampling (h)

 

 

Sex

 

No. of surviving dosed mice

Erythrocytes

Polychromatic cells (PCE)

 

Mean PCE/NCE

 

 

PCE Analysed

No. of MN-PCE

% MN-PCE

Negative control

0+24+48

72

M/F

10

10000

14

0.14

0.93

96

M/F

10

10000

10

0.10

1.02

Positive control

0+24+48

72

M/F

10

10000

150*

1.50

0.46

500

0+24+48

72

M/F

10

10000

9

0.09

1.00

1000

0+24+48

72

M/F

10

10000

9

0.09

0.94

2000

0+24+48

72

M/F

10

10000

8

0.08

0.86

96

M/F

10

10000

24

0.24

0.90

PCE = Polychromatic erythrocytes                                                           

MN-PCE = Micronucleated PCE

MN-PCE = Micronucleated NCE

* = Positive response in PCE

Conclusions:
Fatty alcohol blend has been tested according to OECD 474 and under GLP. Male and female mice were dosed with 500, 1000 and 2000 mg/kg bw. No increase in the number of micronucleated PCE was observed (1000 PCE scored per animal). It is concluded that the test substance is negative for the induction of micronuclei under the conditions of the test. No toxicity to bone marrow or general toxicity was observed.
Executive summary:

Fatty alcohol blend has been tested according to OECD 474 and under GLP. Male and female mice were dosed with 500, 1000 and 2000 mg/kg bw. No increase in the number of micronucleated PCE was observed (1000 PCE scored per animal). It is concluded that the test substance is negative for the induction of micronuclei under the conditions of the test. No toxicity to bone marrow or general toxicity was observed.

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
not stated
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Only 1000 erythrocytes were scored per animal, full experimental details were not reported, toxicity details were lacking. It was not compliant with GLP.
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
(only 1000 PCEs per animal scored for micronuclei)
Principles of method if other than guideline:
Well-conducted study according to protocol very similar to OECD guideline 474
GLP compliance:
not specified
Type of assay:
micronucleus assay
Species:
mouse
Strain:
NMRI
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: BRL Tierfarm Fullinsdorf, Switzerland
- Age at study initiation: >=10 weeks
- Weight at study initiation: no data
- Assigned to test groups randomly: no data
- Fasting period before study: 18 hours, but continued to receive water ad libitum
- Housing: Markrolon Type 1 cages with wire mesh tops and granulated soft wood bedding
- Diet (e.g. ad libitum): standard pellet diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21
- Humidity (%): not regulated
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: no data
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: polyethylene glycol
- Justification for choice of solvent/vehicle: no data
- Concentration of test material in vehicle: no data [calculated: 5, 15 and 50 mg/ml]
- Amount of vehicle (if gavage or dermal): 10 ml/kg bw
- Lot/batch no. (if required): no data
- Purity: no data
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: few details; test material suspended in vehicle
Duration of treatment / exposure:
single administration
Frequency of treatment:
single administration
Post exposure period:
none
Dose / conc.:
50 mg/kg bw/day (nominal)
Dose / conc.:
150 mg/kg bw/day (nominal)
Dose / conc.:
500 mg/kg bw/day (nominal)
No. of animals per sex per dose:
6
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide
- Justification for choice of positive control(s): no data
- Route of administration: presumably oral gavage
- Doses / concentrations: 40 mg/kg bw
Tissues and cell types examined:
bone marrow cells
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: based on previous study - 500 mg/kg bw estimated to be the "maximum attainable dose"
TREATMENT AND SAMPLING TIMES (in addition to information in specific fields): 24, 48 and 72 hours after dosing

DETAILS OF SLIDE PREPARATION: femurs removed, marrow flushed out with foetal calf serum, cell suspension centrifuged and supernatant discarded, small drop of cell pellet spread on slide, air dried, stained with May-Grunwald, mounted; 1 slide/sample

METHOD OF ANALYSIS: 1000 polychromatic erythrocytes (PCEs) scored for micronuclei; polychromatic:normochromatic (PCE:NCE) ratio scored

OTHER: only 5/sex per dose level evaluated
Evaluation criteria:
To be considered positive, either a statistically significant dose-related increase in the number of micronucleated PCEs or a reproducible, statistically significant positive response for at least one dose level
Statistics:
Mann-Whitney test
Sex:
male/female
Genotoxicity:
negative
Toxicity:
not specified
Remarks:
presumably toxic at >500 mg/kg bw
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: no data
- Solubility: no data
- Clinical signs of toxicity in test animals: no data
- Evidence of cytotoxicity in tissue analyzed: no data
- Rationale for exposure: no data
- Harvest times: no data
- High dose with and without activation: no data
- Other: presumably toxic above 500 mg/kg bw since this maximum dose was chosen for the main study on the basis of the results of the range-finding study

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): 0.03-0.09% for vehicle controls, 0.04-0.10% for test material treated, 0.71% for positive control
- Ratio of PCE/NCE (for Micronucleus assay): 1.05-1.27 for vehicle controls, 0.98-1.55 for test material treated, 0.93 for positive control
- Appropriateness of dose levels and route: appropriate (top dose was apparently the maximum tolerated dose, oral route relevant to humans)
- Statistical evaluation: no statistically significant increases in the frequency of micronuclei in mice treated with the test material; statistical significance not presented for positive control

Toxicity unclear, but possibly one male and one female mouse [per group?] died either spontaneously or due to gavage error.

Table 1 Results of micronucleus assay 24 hour sampling time

Treatment

Suspending agent

Low dose

Mid dose

High dose

Concentration mg/kg bw

0

40

50

150

Harvest time

24

24

24

24

Micronucleated PCE (%)

0.03

0.71

0.07

0.08

Ratio PCE/NCE

1.27

0.93

0.98

1.07

Table 2 Results of micronucleus assay 48 hour sampling time

Treatment

Suspending agent

Test substance

Test substance

Test substance

Concentration mg/kg bw

0

50

150

500

Harvest time

48

48

48

48

Micronucleated PCE (%)

0.09

0.1

0.04

0.05

Ratio PCE/NCE

1.05

1.06

1.01

1.23

Table 3 Results of micronucleus assay 72 hour sampling time

Treatment

Suspending agent

Low dose

Mid dose

High dose

Concentration mg/kg bw

0

50

150

500

Harvest time

72

72

72

72

Micronucleated PCE (%)

0.09

0.09

0.05

0.07

Ratio PCE/NCE

1.41

1.33

1.55

1.46

Conclusions:
In a reliable study, behenyl alcohol (C22) did not increase the incidence of micronuclei in mouse bone marrow cells after a single oral gavage dose of up to 500 mg/kg bw.
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
11-Feb-1992 to 27-Apr-1992
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
micronucleus assay
Species:
mouse
Strain:
other: albino mice, CFW 1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Winkelmann
- Age at study initiation: 7-8 weeks
- Weight at study initiation: males 21-27 g, females 21-26 g
- Assigned to test groups randomly: yes, under following basis: allocated to treatment groups according to randomization table generated by computer programme or manually
- Fasting period before study: yes, overnight until 3-4 hours after dosing
- Housing: males, 1/cage, macrolon cages type I; females, <=3/cage, macrolon cages type II; filled with clean softwood bedding
- Diet (e.g. ad libitum): standard animal diet, Altromin No. 1314, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: >=6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25 +- 3 (occasionally 20-25)
- Humidity (%): 40 - 50 (occasionally 45-70)
- Air changes (per hr): no data, except "air-conditioned room"
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES (main study): From: 25-Feb-1992 To: 28-Feb-1992
Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: arachis oil
- Justification for choice of solvent/vehicle: test material easily soluble at required concentration
- Concentration of test material in vehicle: not stated, but provided a dose level of 5000 mg/kg bw, so 500 mg/ml
- Amount of vehicle (if gavage or dermal): 10 ml/kg bw (main study), 20 ml/kg bw (range finding study)
- Lot/batch no. (if required): no data
- Purity: no data
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: 500 mg/ml in arachis oil (main study)
Duration of treatment / exposure:
single administration
Frequency of treatment:
single administration
Post exposure period:
evaluated at 24, 48, 72 hours after administration
Dose / conc.:
5 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
6
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide
- Justification for choice of positive control(s): not stated
- Route of administration: oral
- Dose: 20 mg/kg bw
Tissues and cell types examined:
bone marrow
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: maximum tolerated dose, based on range-finding study (effects seen at 5000 mg/kg were piloerection only)

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): single administration, animals sacrificed 24, 48 and 72 hours after administration

DETAILS OF SLIDE PREPARATION: bone marrow collected from femurs, using foetal calf serum applied via a syringe, into a siliconised centrifuge tube; after centrifugation at 1000 rpm and removal of all but one drop of supernatant, cells of sediment carefully mixed; drop of cell suspension placed on clean, degreased microscope slide and immediately spread; 3 slides/animal; slides air dried at least overnight; stained with Giemsa; air dried and dipped in xylol for 3 minutes

METHOD OF ANALYSIS: 1 slide/animal chosen and given a random code; microscopic evaluation of slides from 5 males and 5 females per treatment group at 1000x magnification; number of micronucleated cells counted in 1000 polychromatic erythrocytes (PCEs)/animal; ratio of
polychromatic to normochromatic erythrocytes determined by counting and differentiating the first 1000 erythrocytes

OTHER: means and standard deviations calculated
Evaluation criteria:
Statistically significant (p<0.05) increase in PCE compared to controls at any sampling time in either sex
Acceptability of test: positive controls induced statistically significant increase in frequency of micronucleated PCEs; solvent control incidence of micronuclei should reasonably fall within historical control range for the testing facility.
Statistics:
Method used: Kastenbaum & Bowman
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Remarks:
piloerection for 8 hours after administration; no mortality
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: 5000 mg/kg bw
- Solubility: used at 250 mg/ml
- Clinical signs of toxicity in test animals: piloerection
- Evidence of cytotoxicity in tissue analyzed: not examined
- Rationale for exposure: based on limit test in rats in which acute oral LD50 was >5000 mg/kg bw
- Harvest times: animals observed for 3 days

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): no statistically significant increase in micronucleus frequency in any treatment group
- Ratio of PCE/NCE (for Micronucleus assay): treated groups similar to controls
- Appropriateness of dose levels and route: maximum tolerated dose of 5000 mg/kg bw used; guideline recommends maximum dose of 2000 mg/kg bw; oral route selected "taking into account the possible route of human exposure during manufacture, handling and use"
- Statistical evaluation: no statistically significant increases in micronuclei in treated groups of either sex; positive control produced a statistically significant increase in micronuclei
- Control incidence of micronuclei: not reported but presumably therefore within historical control range

Lorol 12 did not increase the frequency of micronucleated erythrocytes or the PCE:NCE ratio in mice at any time interval after treatment (24, 48 or 72 hours) at dose levels up to 5000 mg/kg bw when compared to vehicle controls.

Mean values per group in the micronucleus test with 1-Dodecanol (Lorol C12-99)

a) Number of micronucleated cells per 1000 polychromatic erythrocytes (PCE)

b) Ratio of polychromatic to normochromatic erythrocytes (PCE/NCE)

Treatment group; (sampling time)

Species and sex

Dose mg/kg

Micronucleated cells 1000 PCE

Ratio of PCE/NCE

Mean

Range

Mean

Range

Negative control (24 hours) arachis oil

male mice

10 ml/kg

3.60

0 - 9

1.11

0.80 - 1.31

female mice

10 ml/kg

2.00

0 - 4

1.34

1.02 - 1.07

Positve control (24 hours) cyclophosphamide

male mice

20

13.40

10 - 16

1.21

0.90 - 1.72

female mice

20

10.80

7 - 14

0.95

0.67 - 1.28

1-Dodecanol (Lorol C12-99)

 

 

 

 

 

 

 

Limit dose (24 hours)

male mice

5000

2.60

0 - 5

1.08

0.94 - 1.26

female mice

5000

2.40

2 - 3

1.01

0.90 - 1.18

Limit dose (48 hours)

male mice

5000

3.00

1 - 4

0.89

0.48 - 1.16

female mice

5000

2.00

0 - 5

1.18

0.90 - 1.68

Limit dose (72 hours)

male mice

5000

2.60

2 - 4

1.65

0.91 - 2.14

female mice

5000

1.60

0 - 4

1.33

1.08 - 1.55

 

Conclusions:
Dodecan-1-ol has been tested a reliable study, conducted according to OECD guideline 474, no genotoxicity was seen in mice after a single oral dose of 5000 mg/kg bw. The study was performed in compliance with GLP.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Relaible information on the genetic toxicity of tetradecan-1-ol was available for bacterial mutagenicity. For endpoints where no information was available, key studies were chosen from studies on closely related linear or branched alcohols of similar chain length. The choice of key study was based on reliability and similarity of chain length. The data available from standard in vitro and in vivo genetic toxicity assays for all related substances show no evidence of mutagenic potential.

Discussion of trends in the Category of C6-24 linear and essentially-linear aliphatic alcohols:

Thein vitroandin vivodata available for members of the category and supporting substances indicate that the C6-24 alcohols are not genotoxic. In addition, the category of LCAAs under consideration does not contain any structural elements that are of concern for potential mutagenic activity (Ashby and Tenant, 1991). Furthermore, primary LCAAs (linear and branched) in the range C1 to C5 do not have a mutagenic potential (Bevan, 2001; OECD SIDS butan-1-ol, 2001). Moreover, in a review by WHO-JECFA a series of 22 saturated aliphatic branched-chain primary LCAAs and the corresponding aldehydes and acids in the range C4 to C8 showed no activity in a battery of in vitro and in vivo mutagenicity tests (WHO, 1999). On this basis it is concluded that the category of LCAAs does not have a mutagenic potential and that read-across within the category can be justified. Where data gaps exist, the gap is filled by read-across from reliable evidence within the C6-24 Alcohols Category, where possible using interpolation between at least two reliable studies using higher and lower carbon number test substances.

Conclusion:                                                                                                                                                        

The category C6-24 LCAAs do not have a genotoxic potential.

 Genetic toxicity of LCAAs

 

CAS

CHEMICAL NAME

Bacterial mutagenicity

Bacterial mutagenicity

Mammalian cytogenicity

Mammalian cytogenicity

Mamalian mutagenicity

Mamalian mutagenicity

In vivostudies

In vivostudies

 

 

 

Result (Rel.)

Reference

Result (Rel.)

Reference

Result (Rel.)

Reference

Result (Rel.)

Study Type*(Ref)

C6

111-27-3

Hexan-1-ol

 Neg; (1)

 Henkel, 1990

 

 

 

 

 

 

C7, 8 and 9

 

Alcohols, C7-9

Neg. (1)

Shell, 1996

 

 

 

 

 

 

C8

111-87-5

Octan-1-ol

Neg; (2)

 Henkel, 1982a; HLS, 1996k

 

 

 

 

 

 

C8

104-76-7

2-ethylhexan-1-ol

Supporting Substance

Neg; (2)

Kirby, 1983

 

 

Neg; (2)

Kirby, 1983

Neg; (2)

MN;Dom. Leth

(Putman, 1983; WHO, 1993)

C8-10

none

Fatty alcohol blend (40.7% C8 and 55.3% C10)

Supporting Substance

Neg(2)

Dillon, D.M., McCartney, M.A. (1992)

 

 

Neg (1)

Cattanach, P., Riach, C. (1992)

Neg (1)

Micronucleus (Holstrom, M., Innes, D. (1992))

C10

112-30-1

Decan-1-ol

Neg (4) 2 strains only

 

 (HLS, 1996l)

 

 

 

 

 

 

C12

112-53-8

Dodecan-1-ol

Neg. (1)l

 (Thompson, 1996a)Shimizu, 1985

 

 

 

 

Neg. (2)

Micronucleus; (Banduhn, 1992)

C12 and 13

75782-87-5

Alcohols, C12-13

Neg (2, >80% lin)

 Dean, 1980

 

 

 

 

 

 

C12 and 13

740817-83-8

Alcohols, C12-13-branched and linear

Neg (1 50% lin),

Sasol, 1998f

Neg (1 (50% lin)

Sasol, 1998 g

 

 

 

 

C12

67762-25-8

C12-18 Alcohols, Type B

Supporting

Neg (2)Ames

Henkel 1982c

 

 

 

 

 

 

C 12-15

90604-40-3

Alcohols, C12-15-branched and linear

Neg (1)

Ballantyne, 1996

 

 

 

 

 

 

C14

112-72-1

Tetradecan-1-ol

Neg (1)

Thompson, 1996b

 

 

 

 

 

 

C16

36653-82-4

Hexadecan-1-ol

Neg (1)

Thompson, 1996c

 

 

 

 

 

 

C16

36653-82-4

Hexadecan-1-ol

Neg. (2)

Henkel, 1981d

 

 

 

 

 

 

C16

68002-94-8

C16-18 and C18 Unsaturated

Supporting

Neg. Ames (2)

Banduhn, 1989)

 

 

 

 

 

 

C18

112-92-5

Octadecan-1-ol

Neg (1)

 

Thompson, 1996d

 

 

 

 

 

Neg (2) MN

Hachiya, 1982

C18

112-92-5

Octadecan-1-ol

Neg(2)

Henkel, 1981f

 

 

 

 

 

 

C18

97552-91-5

C18-22 Alcohol

Supporting

Neg. Ames (2)

 Banduhn 1995

 

 

 

 

 

 

C22

661-19-8

Docosan-1-ol

Neg (2),

 

Iglesias, 2002a, Thompson, 1997

Neg (2),

Iglesias, 2002a

Neg (2)

Iglesias, 2002a

Neg (2)

Micronucleus Iglesias, 2002aª

C24-32

 

D-002***

Supporting substance

 

 

 

 

 

 

Neg (4)

MN; Dom. Leth.Rodeiro 1998a

                                                                                                                                                                                        

* MN: Mouse bone marrow micronucleus test; Dom. Leth. Mouse Dominant Lethal test; UDS: Unscheduled DNA Synthesis assay

** Tested in S. typhimurium TA 98 and TA100, only.

***Mixture of very long chain fatty alcohols from hydrolysed beeswax

References:                                                                                                                                                      

Ashby, J., Tennant, R.W., 1991. Definitive relationships among chemical structure, carcinogenicity, and mutagenicity for 301 chemicals tested by the US NTP. Mutation Research 257, 229–306.          

WHO, 1999. Technical Report Series 884 Evaluation of certain food additives and contaminants. 49th Report of the Joint FAO/WHO Expert Committee on Food Additives (JECFA), Geneva.


Justification for classification or non-classification

Based on the available data, tetradecan-1ol does not require classification for genetic toxicity according to Regulation (EC) No 1272/2008.