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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study (OECD 471); GLP compliant.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report date:
2005

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Phthalide
EC Number:
201-744-0
EC Name:
Phthalide
Cas Number:
87-41-2
Molecular formula:
C8H6O2
IUPAC Name:
1(3H)-Isobenzofuranone
Details on test material:
- Name of test material (as cited in study report): Phthalide
- Physical state: Solid, white
- Analytical purity: 100%
- Lot/batch No.: S12233-025
- Stability under test conditions: The stability of the test substance under storage conditions throughout the study period is guaranteed.
- Storage condition of test material: room temperature

Method

Target gene:
his and trp
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
S9-mix from Aroclor 1254 treated male Sprague-Dawley rats.
Test concentrations with justification for top dose:
0; 55; 174; 550; 1740 and 5500 µg/plate (SPT)
0; 55; 174; 550; 1740 and 5500 µg/plate (PIT )
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
Controls
Untreated negative controls:
yes
Remarks:
sterility control; plates are treated with soft agar, S-9 mix, buffer, vehicle or the test substance but without the addition of tester strains.
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: with S9: 2-aminoanthracene; without S9: N-methyl-N'-nitro-N-nitrosoguanidine, 4-nitro-o-phenylendiamine, 9-aminoacridine, 4-nitroquinoline-N-oxide
Details on test system and experimental conditions:
METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 20 min
- Exposure duration: 48 h - 72 h

NUMBER OF REPLICATIONS: 3


METHOD OF APPLICATION: standard plate test

DURATION
- Exposure duration: 48 h - 72 h

NUMBER OF REPLICATIONS: 3
Evaluation criteria:
The test chemical is considered positive in this assay if the following criteria are met: A dose-related and reproducible increase in the number of revertant colonies, i.e. about doubling of the spontaneous mutation rate in at least one tester strain either without S-9 mix or after adding a metabolizing system.

Results and discussion

Test resultsopen allclose all
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:
cytotoxicity
Remarks:
A weak bacteriotoxic effect was occasionally observed only in the preincubation assay at 5500 µg/plate .
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
A weak bacteriotoxic effect was occasionally observed only in the preincubation assay at 5500 µg/plate .
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: precipitation of the test substance was found.

Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Standart plate test:

 Dose (µg/plate)  TA1535     TA100     TA1537     TA98    E. coli                                              
   -S9  +S9  -S9  +S9  -S9  +S9 -S9  +S9 - S9 + S9                                              
0

16±2

17±3

111±8

107±7

9±2

10±2

31±6

33±3

42±1

48±7

 

 

 

 

 

 

                                 

55

18±3

16±3

107±2

110±12

10±10

10±3

30±5

29±2

41±3

38±6

 

 

 

 

 

 

                                 

174

17±2

15±2

108±5

99±8

8±2

10±2

27±4

29±3

43±9

40±6

 

 

 

 

 

 

                                 

550

20±3

15±2

103±6

104±2

8±1

7±1

28±2

39±2

45±2

42±4

 

 

 

 

 

 

                                 

1740

15±4

18±3

102±4

106±4

11±4

7±1

27±4

41±7

41±5

48±9

 

 

 

 

 

 

                                 

5500

13±3

15±3

108±8

106±6

6±3

5±3

27±3

25±4

38±8

40±1

 

 

 

 

 

 

                                 

2AA

-

126±11

-

820±38

-

115±6

-

642±34

-

267±31

 

 

 

 

 

 

                                 

MNNG

654±27

-

800±44

-

-

-

-

-

-

-

 

 

 

 

 

 

                                 

AAC

-

-

-

-

416±27

-

-

-

-

-

 

 

 

 

 

 

                                 

NOPD

-

-

-

-

-

-

493±62

-

-

-

 

 

 

 

 

 

                                 
4NQO  -  -  -  -  -  -  -  -  1256±31  -                                              

Mean ± SD

Preincubation-test:

 Dose (µg/plate)  TA1535     TA100     TA1537     TA98    E. coli                                              
   -S9  +S9  -S9  +S9  -S9  +S9 -S9  +S9 - S9 + S9                                              
0

18±2

17±3

109±5

110±6

10±2

10±2

29±3

34±6

32±4

34±5

 

 

 

 

 

 

                                 

55

15±3

17±3

109±5

104±5

9±2

8±2

26±3

30±3

28±7

31±4

 

 

 

 

 

 

                                 

174

18±1

15±1

114±7

104±17

8±0

8±2

31±3

33±13

29±11

35±8

 

 

 

 

 

 

                                 

550

14±1

13±4

107±13

105±9

7±3

8±2

23±2

28±6

27±6

25±2

 

 

 

 

 

 

                                 

1740

15±4

16±4

99±9

102±2

7±1

6±2

27±4

26±4

23±3

25±4

 

 

 

 

 

 

                                 

5500

16±2

10±1

74±6

79±14

4±1

4±1

21±3

24±4

24±4

24±5

 

 

 

 

 

 

                                 

2AA

-

115±15

-

644±58

-

109±10

-

229±13

-

215±7

 

 

 

 

 

 

                                 

MNNG

638±44

-

712±54

-

-

-

-

-

-

-

 

 

 

 

 

 

                                 

AAC

-

-

-

-

439±10

-

-

-

-

-

 

 

 

 

 

 

                                 

NOPD

-

-

-

-

-

-

461±45

-

-

-

 

 

 

 

 

 

                                 
4 NQO  -  -  -  -  -  -  -  -  536±19  -                                              

Mean ± SD

X: reduced background growth

2-AA: 2-aminoanthracene;

MNNG; N-methyl-N-nitro-N-nitrosoguanidine

NOPD: 4-nitro-o-phenylendiamine

AAC: 9-aminoacridine chloride monohydrate

4-NQO: 4-nitroquinoline-N-oxide

 

 

According to the results of the present study, the test substance Phthalide is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay under the experimental conditions chosen here. The positive controls gave the expected values.

Applicant's summary and conclusion