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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
Study period:
2002-03-13 to 2002-03-28
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- name of testsubstance used in report: Praepagen HY
- Physical state: yellowish, liquid
- Batch No.: ESDB 01 496
- active ingrdient /purity: Fattyalkyl dimethylhydroxyethylammonium chloride/ ca. 40% in water
- Expiration date of the lot/batch: 2003-07-19
- Storage condition of test material: room temperature (20°C)
- pH: 5.5 - 8.5 (20°C, 10g/L)

Method

Species / strainopen allclose all
Species / strain:
S. typhimurium TA 100
Details on mammalian cell lines (if applicable):
- Properly maintained: yes
- Periodically checked for karyotype stability: yes
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Postmitochondrial (S9) fraction from livers of male wistar rats, induced with Phenobarbital (i.p.) and ß-Naphtoflavon orally.
Species / strain:
S. typhimurium TA 98
Details on mammalian cell lines (if applicable):
- Properly maintained: yes
- Periodically checked for karyotype stability: yes
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Postmitochondrial (S9) fraction from livers of male wistar rats, induced with Phenobarbital (i.p.) and ß-Naphtoflavon orally.
Species / strain:
S. typhimurium TA 97
Details on mammalian cell lines (if applicable):
- Properly maintained: yes
- Periodically checked for karyotype stability: yes
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Postmitochondrial (S9) fraction from livers of male wistar rats, induced with Phenobarbital (i.p.) and ß-Naphtoflavon orally.
Species / strain:
S. typhimurium TA 102
Details on mammalian cell lines (if applicable):
- Properly maintained: yes
- Periodically checked for karyotype stability: yes
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Postmitochondrial (S9) fraction from livers of male wistar rats, induced with Phenobarbital (i.p.) and ß-Naphtoflavon orally.
Species / strain:
S. typhimurium TA 1535
Details on mammalian cell lines (if applicable):
- Properly maintained: yes
- Periodically checked for karyotype stability: yes
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Postmitochondrial (S9) fraction from livers of male wistar rats, induced with Phenobarbital (i.p.) and ß-Naphtoflavon orally.
Test concentrations with justification for top dose:
1. Study:
all strains (±S9) : 0.005- 0.016 - 0.05 - 0.16 - 0.5 (mg/plate)
2. Study:
TA 97a, TA 100 (±S9), TA 98, TA 102, TA 1535 (-S9): 0.0016 - 0.005- 0.016 - 0.05 - 0.16 (mg/plate)
TA 98, TA 102, TA 1535 (+S9): 0.016 - 0.05 - 0.16 - 0.5 - 1.6 (mg/plate)
Negative controls tested with aqua bidest.
Vehicle:
aqua bidest
Controlsopen allclose all
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: ICR 191
Remarks:
positive control in TA 97a (-S9)
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2- Aminoanthracen
Remarks:
positive control in TA 97a, TA 98, TA 100, TA 1535 (+S9)
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
cumene hydroperoxide
Remarks:
positive control in TA 102 (–S9)
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 4-nitro-o-phenylendiamine
Remarks:
positive control in TA 98 (– S9)
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Nitrofurantoine
Remarks:
positive control in TA 100 (– S9)
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Sodiumazide
Remarks:
positive control inTA 1535 (– S9)
Negative controls:
yes
Solvent controls:
yes
Remarks:
aqua bidest
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Danthron
Remarks:
positive control in TA 102 (+S9)
Details on test system and conditions:
METHOD OF APPLICATION: in agar (plate incorporation);
- Two independent studies were conducted
- Exposure duration: 48h
- NUMBER OF REPLICATIONS: 3 replicates per concentration level and control
Evaluation criteria:
Reduced background lawn is regarded to be a cytotoxic effect. Plates with reduced background lawn were not included into evaluation.
Mean values and standard deviations were calculated from colonies per plate of three replicates.
Induction rate = revertant colonies of test item divided by revertant colonies of the control.
The test item is to be interpretated mutagenic if there is a concentration effect relationship and the induction rate is ≥ 2.
Statistics:
The arithmetic mean value and the standard deviation were calculated out of three replicates of the revertant colony plates

Results and discussion

Test results
Species / strain:
other: TA 97, TA 98, TA 100, TA 1535, TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
yes
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
Based on the results of a preliminary test (non GLP) the test concentrations have been selected.

Any other information on results incl. tables

Table 1: Mutagenic and Cytotoxic Effects of the Test Item Praepagen HY

Strain

S9

TestedConcentrationRange[mg/plate]

Lowest Mutagenic Concentration [mg/plate]

Lowest Cytotoxic Concentration [mg/plate]

TA 97a

-

0.0016 – 0.5

none

0.5

+

0.0016 – 0.5

none

0.16

TA 98

-

0.0016 – 0.5

none

0.5

+

0.005 – 1.6

none

1.6

TA 100

-

0.0016 – 0.5

none

0.5

+

0.0016 – 0.5

none

0.5

TA 102

-

0.0016 – 0.5

none

0.16

+

0.005 – 1.6

none

1.6

TA 1535

-

0.0016 – 0.5

none

0.5

+

0.005 – 1.6

none

1.6

 

  Table 2: Summary of mean number of revertants (mean of 3 plates) in Salmonella typhimurium strains with and without metabolic activation

Experiment I

Concentration

TA1535

TA102

TA100

TA98

TA97a

 [mg/plate]

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

Control

24

12

322

382

123

133

22

24

234

275

0.5

n.c.

10

n.c.

353

n.c.

n.c.

n.c.

29.

n.c.

n.c.

0.16

22

13

333

450

74

101

40

40

211

238

0.05

30

15

325

480

126

113

45

35

278

276

0.016

29

15

397

457

131

100

42

35

277

259

0.005

29

13

344

461

133

100

32

34

263

233

Experiment II

 

TA1535

TA102

TA100

TA98

TA97a

 

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

Control

25

13

389

501

117

112

25

30

260

277

1.6

n.c.

n.c.

0.5

10

203

0.16

17

10

198

346

57

129

28

n.c.

100

n.c.

0.05

12

11

315

333

53

102

31

15

219

121

0.016

25

14

273

364

122

110

34

33

232

226

0.005

16

218

72

100

21

35

231

205

0.0016

30

339

115

93

28

32

116

136

n.c.= not counted, due to reduced background lawn; — = concentration not tested

Applicant's summary and conclusion

Conclusions:
Based on the results of this study the test item was found to have no mutagenic effects on Salmonella typhimurium strains TA 97a, TA 100, TA 98, TA 102, TA 1535 with (+) and without (-) the metabolic activation system S9 from Wistar rats at concentrations up to 0.5mg/plate.