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Toxicological information

Genetic toxicity: in vitro

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Administrative data

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

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Responses of the L1578Y tk+/tk- mouse lymphoma cell forward mutation assay. II. 18 coded chemicals.
Author:
McGregor DB, Brown A, Cattanach P, Edwards I, McBride D, Caspary WJ
Year:
1988
Bibliographic source:
Environ Molec Mutagen 11, 91 - 118
Reference Type:
study report
Title:
Unnamed
Year:
1989
Report date:
1989

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
GLP compliance:
no
Type of assay:
mammalian cell gene mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Hydroquinone
EC Number:
204-617-8
EC Name:
Hydroquinone
Cas Number:
123-31-9
Molecular formula:
C6H6O2
IUPAC Name:
hydroquinone
Details on test material:
- Name of test material (as cited in study report): hydroquinone

Method

Target gene:
Thymidine kinase locus
Species / strain
Species / strain / cell type:
mouse lymphoma L5178Y cells
Details on mammalian cell type (if applicable):
- Type and identity of media: Fischer's medium with 5% heat-inactivated horse serum
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically "cleansed" against high spontaneous background: yes
Metabolic activation:
with and without
Metabolic activation system:
S9 mix from Aroclor 1254 induced rat liver
Test concentrations with justification for top dose:
- S9: 1.56, 3.12, 6.25, 12.5, 25, 50 µg/mL
+ S9: 0.652, 1.25, 2.5, 5, 10 µg/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO (+ S9), methanol (- S9)
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: - S9: EMS; + S9: EMS or 3-methylcholanthrene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 4 h
- Expression time (cells in growth medium): 2 d
- Selection time (if incubation with a selection agent): 11-14 d
- Fixation time (start of exposure up to fixation or harvest of cells): 13-16 d

SELECTION AGENT (mutation assays): trifluorothymidine

NUMBER OF REPLICATIONS: 2, two independent assays

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth
Evaluation criteria:
Positive response: dose-related trend and statistically significant response at one of the three highest doses
Statistics:
Dose trend test (Barlow et al., 1972); variance analysis (pairwise comparisons) with P < 0.05

Results and discussion

Test results
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

Table: Results of the mouse lymphoma cell mutation assay without and with metabolic activation

Metabolic activation / trial

Test concentration

Cloning efficiency (%)

Relative total growth (%)

Mutant fraction

Average mutant fraction

- S9 / trial 1

0

47

68

73

74

35

57

96

48

164

62

60

110

65

1.562

41

45

200

157*

42

108

113

3.125

25

23

454

333*

42

50

213

6.25

17

7

857

624*

25

13

391

12.5

TOX

TOX

Positive control

47

101

214

197*

45

43

181

- S9 / trial 2

0

76

96

19

22

61

100

28

93

98

18

85

106

24

3.125

13

5

565

419*

57

30

274

6.25

48

13

542

542*

37

9

541

12.5

27

7

871

857*

25

7

843

25

28

6

863

774*

30

7

685

50

TOX

TOX

Positive control

120

114

108

108

- S9 / trial 3

0

93

114

19

38

62

109

47

52

90

42

90

87

45

0.625

81

68

47

49

102

71

51

1.25

36

22

164

179*

27

16

195

2.5

15

7

607

624*

12

4

642

5

TOX

12

1

1915

10

TOX

TOX

Positive control

69

66

299

306*

93

58

313

+ S9 / trial 1

0

91

99

65

70

70

102

67

68

98

70

79

100

77

0.625

93

116

67

72

90

133

78

1.25

81

105

69

78

72

99

86

2.5

75

81

138

134*

98

93

130

5

61

20

338

328*

70

18

318

10

43

7

485

464*

44

7

582

Positive control

55

57

423

451*

45

47

480

* Statistically significant increase compared to vehicle controls, P < 0.05

Applicant's summary and conclusion

Conclusions:
Positive with and without metabolic activation.

In the mouse lymphoma assay for induction of trifluorothymidine resistance in L5178Y/TK cells, HQ was positive at doses of 1.25 µg/mL and higher in the absence of S9, and at 2.5 µg/mL and higher in the presence of S9.
Executive summary:

Hydroquinone was tested in a mouse lymphoma mutation assay for induction of trifluorothymidine resistance in L5178Y/TK cells with a test protocol similar to OECD Guideline 476. Test concentrations of HQ ranged from 0.625-50 µg/mL without S9 mix and 0.625-10 µg/mL with S9 mix. Treatment time was 4 h, followed by a 2-day expression period and a 11 to 14-day selection period in the presence of TFT. Cytotoxicity was indicated by relative total growth. The lowest effective dose with a significantly increased mutant fraction was 1.25 µg/mL without S9 (relative total growth 32%), and 2.5 µg/mL with S9 (relative total growth 87%). The mutant fraction was statistically significantly and dose-dependantly increased from these concentrations up. HQ was positive in the mouse lymphoma assay both in the presence and absence of S9.