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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1994-04-06 t 1994-06-01
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1994
Report date:
1994

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
Version / remarks:
Standards for Toxicity Investigations (Japan's Ministry of Labor, No. 77, September 1, 1988) and Notification on Partial Revision of Testing Methods Relating to the New Chemical Susbtances (Notification No. 700 of the Planning and Coordination Bureau, EA, No. 1039 of the Pharmaceutical Affairs Bureau, MHW & No. 1014 (1986) of the Basic Industries Bureau, MITI, December 5, 1986).
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Trideca-1,1,1,2,2,3,3,4,4,5,5,6,6-fluorohexane
EC Number:
206-581-9
EC Name:
Trideca-1,1,1,2,2,3,3,4,4,5,5,6,6-fluorohexane
Cas Number:
355-37-3
Molecular formula:
C6HF13
IUPAC Name:
Trideca-1,1,1,2,2,3,3,4,4,5,5,6,6-fluorohexane
Test material form:
liquid
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Asahi Glass Co., Ltd.; Batch no. 31001
- Expiration date of the lot/batch: Not specified
- Purity test date: Not specified

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Stored in a cold and dark place
- Stability under test conditions: Not specified

FORM AS APPLIED IN THE TEST (if different from that of starting material): Colourless clear liquid

OTHER SPECIFICS:

Purity: 99.9%
Impurities: CClF2HCCl2F (0.05%)
Molecular Weight: 320.05
Boiling Point 71°C
Solubility: Oil soluble
Degree of Solublity Water (<10 mg/L)
DMSO (<5% w/v)
Acetone (≥ 10% w/v)

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Rat Liver S9
Test concentrations with justification for top dose:
Dose range-finding test: 0, 50, 100, 200, 500, 1000, 2000, or 5000 µg/plate

Main Test: 0, 313, 625, 1250, 2500, or 5000 µg/plate (Based on the results of the dose range-finding test, 5000 µg/plate was selected as the highest dose and the lower four doses diluted with a geometric progression of 2 were set).
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: Not specified
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
not specified
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
sodium azide
other: 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2); 2-Aminoanthracene (2AA); 2-Methoxy-6-chloro--9-[3-(2-chloroethyl)-aminopropylamino]acridine.2HCl
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)
- Cell density at seeding (if applicable): Main Test - TA98: 2.0 x 10^9 viable cells/mL; TA100: 1.6 x 10^9 viable cells/mL; TA1535: 2.1 x 10^9 viable cells/mL; TA1537: 1.8 x 10^9 viable cells/mL; WP2 uvrA: 1.9 x 10^9 viable cells/mL

DURATION
- Preincubation period: 37 ± 0.5°C for 8-10 hours
- Expression time (cells in growth medium): 20 minutes
- Exposure duration: 48 hours

SELECTION AGENT (mutation assays): Not specified

NUMBER OF REPLICATIONS: 3 (negative control); 2 (Test substance and positive controls)

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: The appearance of revertant colonies (size and number), deposition of the test substance and growth inhibition were examined with a stereo microscope.

NUMBER OF CELLS EVALUATED: The number of revertant colonies was counted with a manual counter or a colony analyzer (CA-7, Toyo-sokki Co., Ltd.). The count using the colony analyzer was finalized after correcting with count drop. Each plate was measured three times, and the average of these three individual measurements with rounding off the figures below decimal point was adopted as the number of revertant colonies.
Evaluation criteria:
The test substance was judged to be positive, when the number of revertant colonies was increased more than twice that of the negative control in a dose-dependant manner, and the reproducibility of results was also obtained. It was judged to be negative in the other cases.
Statistics:
Statistical analysis was not applied.

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
not examined
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
Vehicle controls validity:
not examined
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
not examined
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
Vehicle controls validity:
not examined
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
not examined
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
The sterility test confirmed the absence of micro-organisms.

Any other information on results incl. tables

Table 1. Results of the Main Test

With (+)

or Without

(-S9) Mix

Test Substance Dose

(µg/plate)

Number of Revertants per Plate

 

Base-pair Substitution Type

 

Frameshift Type

TA 100

TA 1535

WP2uvrA

TA 98

TA 1537

-S9 Mix

Negative Control

131

153 (134)

117

12

8 (10)

9

44

47 (43)

37

22

20 (19)

14

10

12 (10)

8

313

174

152 (163)

17

13 (15)

61

46 (54)

26

34 (30)

15

4 (10)

625

119

113 (116)

9

18 (14)

52

53 (53)

17

24 (21)

22

5 (14)

1250

162

158 (160)

14

15 (15)

51

43 (47)

24

28 (26)

5

11 (8)

2500

147

147 (147)

10

17 (14)

48

42 (45)

19

29 (24)

11

12 (12)

5000

147

147 (147)

11

15 (13)

41

43 (42)

26

16 (21)

8

7 (8)

 

+S9 Mix

Negative Control

148

148 (148)

148

21

28 (22)

16

51

47 (49)

49

34

29 (34)

39

24

19 (17)

8

313

149

168 (159)

13

18 (16)

57

52 (55)

41

37 (39)

14

22 (18)

625

133

143 (138)

11

14 (13)

50

61 (56)

43

48 (46)

22

13 (18)

1250

147

134 (141)

14

12 (13)

53

52 (53)

29

30 (30)

11

25 (18)

2500

112

147 (130)

12

14 (13)

55

41 (48)

38

30 (34)

17

19 (18)

5000

158

138 (148)

15

22 (19)

54

56 (55)

36

35 (36)

15

20 (18)

 

Positive Control

(-S9 Mix)

Chemical

AF-2

NaN3

AF-2

AF-2

ICR-191

Dose

(µg/plate)

0.01

0.5

0.01

0.1

1

Number of Revertants

/plate

424

398 (411)

295

297 (296)

444

412 (428)

395

445 (420)

1925

1948 (1937)

 

Positive Control

(+S9 Mix)

Chemical

2AA

2AA

2AA

2AA

2AA

Dose

(µg/plate)

1

2

10

0.5

2

Number of Revertants

/plate

647

632 (640)

151

130 (141)

447

450 (449)

140

149 (145)

122

123 (123)

1) values in parenthesis show the mean of each plate

2) AF-2: 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2)

3) 2AA: 2-Aminoanthracene

4) ICR-191: 2-Methoxy-6-chloro--9-[3-(2-chloroethyl)-aminopropylamino]acridine.2HCl

5) NaN3: Sodium Azide

Applicant's summary and conclusion

Conclusions:
1,1,1,2,2,3,3,4,4,5,5,6,6-tridecafluorohexane was considered to have no reverse mutagenic potential under the conditions of the study.
Executive summary:

In a key bacterial reverse mutation assay, the in vitro mutagenic potential of the test material, 1,1,1,2,2,3,3,4,4,5,5,6,6-tridecafluorohexane, was evaluated in Salmonella typhimurium strains TA98, TA100, T1535, and TA 1537 and Escherischia coli strain WP2uvrA using pre-incubation method in the absence and presence of a metabolic activation system (±S9).

 

Based on a dose range-finding study, the concentrations of the test material used in the main study were 0, 313, 625, 1250, 2500, or 5000 µg/plate.

 

It was observed that the number of revertant colonies in all strains was less than twice that of the negative control at all dose levels tested in the absence and presence of metabolic activation (±S9). The positive controls exhibited a significant increase in the number of revertant colonies and the number of revertants for positive as well as negative controls was within the historical range.

 

Based on the results observed in the study, 1,1,1,2,2,3,3,4,4,5,5,6,6-tridecafluorohexane was considered to be negative in this bacterial reverse mutation assay.