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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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: - according to OECD 471 as at 1982 - lack of one of the following strains: E. coli WP2 uvrA, or E. coli WP2 uvrA (pKM101), or S. typhimurium TA102. - study only available as summary from the NTP report

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1993
Reference Type:
publication
Title:
Salmonella mutagenicity test results for 250 chemicals.
Author:
Haworth S, Lawlor T, Mortelmans K, Speck W, Zeiger E.
Year:
1983
Bibliographic source:
Environ Mutagen. 1983;5 Suppl 1:1-142. PMID: 6365529

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
as at 1981
Deviations:
no
Principles of method if other than guideline:
.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 1,2,3-Trichloropropane
- Physical state: not reported
- Analytical purity: 99.1% (Midwest Research Institute, MRI)
- Impurities (identity and concentrations): not reported
- Lot/batch No.: JG 32449
- Provider: Shell Chemical (via Radian Corporation (Austin, TX, U.S.A.) as general chemical repository for the NTP program)
- Stability under test conditions: not reported, probably stable
- Storage condition of test material: not reported

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
S9 mix (Aroclor 1254-induced male Sprague-Dawley rat or Syrian hamster liver)
Test concentrations with justification for top dose:
0, 1, 3, 10, 33, 100, 333 µg/plate SRI, International
0, 10, 33, 100, 333, 666, 1'000 µ g/plate Microbial Associates
Vehicle / solvent:
not reported
- Vehicle(s)/solvent(s) used: DMSO; ethanol; or acetone. Not detailed for 1,2,3-Trichloropropane. Probably DMSO was used
- Justification for choice of solvent/vehicle: none
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
other: all strains with rat or hamster S9: 2-Aminoanthracene (2-AA); without S9: TA98 - 4-Nitro-o-phenylenediamine (NOPD), TA100 and TA1535 - sodium azide (SA), TA1537 - 9-aminoacridine. (9-AAD) was tested on TA 1537. The
Details on test system and experimental conditions:
METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 20 min
- Exposure duration: 20 min preincubation + 48 after plating
- Expression time (cells in growth medium): 20 min
- Selection time (if incubation with a selection agent): 48 h


SELECTION AGENT (mutation assays):
minimal glucose bottom agar (lacking histidine; Vogel HJ, Bonner DM (1956): Acetylornithinase of E coil: Partial purification and some properties. J Biol Chem 218:97-106.)

NUMBER OF REPLICATIONS: each concentration and strain in triplicate, test as a whole in duclicate

DETERMINATION OF CYTOTOXICITY
- Method: test with TA 100 at 10 mg/plate, parameters: viability on complete medium (EGG) and reduced numbers of revertant colonies per plate and/or thinning or absence of the bacterial lawn
Evaluation criteria:
- first assessment: reproducible, dose-related increase, whether it be twofold over background or not.
- second assessment: statistical analysis based on Margolin 1981 (Margolin BH, Kaplan N. Zeiger E (1981): Statistical analysis of the Ames Salmonella/microsome test.)
Proc Natl Acad Sci USA 78:3779-3783.
Statistics:
see above

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 97
Metabolic activation:
with
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
other: toxicity starting at 333 µg/Plate with activation and at 666 µg without activation
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
other: SRI: no cytotoxicity up to limit dose 333 µg/plate, which was set based on the sensitivity of TA 100; Microbiological Associates:
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
other: SRI: no cytotoxicity up to limit dose of 333 µg/plate, which was set based on the sensitivity of TA 100; Microbiological Associates: toxic between 666 and 1000 µg/plate, with and without activation
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
other: SRI: no cytotoxicity up to limit dose of 333 µg/plate, which was set based on the sensitivity of TA 100; Microbiological Associates: toxic between 666 and 1000 µg/plate, with and without activation
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: no cytotoxicity up to limit dose of 333 µg/plate, which was set based on the sensitivity of TA 100
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
for details see Tables 1 and 2 below
Remarks on result:
other:
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

- Table 1: Mutagenicity of 1,2,3-Trichloropropane in Salmonella typhimurium performed at SRI, International

Dose µg/plate

Revertants/plate*

-S9

+10% hamster S9

+10% rat S9

Trial 1 

Trial 1 

 Trial 2

Trial 1 

Trial 2 

Strain TA 100

 

 

 

 

 

0

138 ± 11.8

179 ± 9.9

144 ± 4.7

158 ± 6.2

133 ± 4.3

3

145 ± 21.0

267 ± 59.4

210 ± 26.1

141 ± 17.2

130 ± 1.9

10

139 ± 5.6

458 ± 23.9

339 ± 18.6

180 ± 5.3

140 ± 6.5

33

142 ± 14.6

492 ± 75.5

690 ± 24.3

211 ± 16.9

166 ± 9.4

100

135 ± 22.0

816 ± 121.4

1,210 ± 44.4

344 ± 9.8

282 ± 12.8

333

140 ± 7.0

1,005 ± 30.9

1,862 ± 50.8

652 ± 28.6

461 ± 37.9

Trial summary

Negative

Positive

Positive

Positive

Positive

Positive control

352 ± 12.7

2,409 ± 23.4

1,121 ± 67.6

1,079 ± 36.4

688 ± 12.7

Strain TA1535

 

 

 

 

 

0

12 ± 4.1

13 ± 0.0

10 ± 2.6

9 ± 2.7

5 ± 1.0

1

 

 

41 ± 6.1

 

 

3

7 ± 0.9

47 ± 4.4

71 ± 10.0

10 ± 2.6

8 ± 0.9

10

9 ± 1.5

98 ± 18.2

128 ± 20.5

11 ± 3.1

7 ± 1.2

33

7 ± 1.5

209 ± 31.7

266 ± 46.1

31 ± 2.6

21 ± 4.8

100

13 ± 0.6

422 ± 34.6

481 ± 44.6

73 ± 3.5

45 ± 7.9

333

9 ± 0.3

734 ± 109.3

 

205 ± 7.0

80 ± 7.2

Trial summary

Negative

Positive

Positive

Positive

Positive

Positive control

294 ± 30.5

514 ± 7.3

179 ± 5.7

225 ± 18.5

103 ± 14.3

Strain TA1537

 

 

 

 

 

0

5 ± 2.2

6 ± 0.6

 

6 ± 2.1

 

3

4 ± 0.9

7 ± 1.3

 

4 ± 0.9

 

10

4 ± 0.7

8 ± 0.3

 

4 ± 0.6

 

33

5 ± 1.8

8 ± 0.0

 

5 ± 1.0

 

100

6 ± 1.3

12 ± 2.4

 

6 ± 0.9

 

333

5 ± 1.3

7 ± 3.2

 

10 ± 2.2

 

Trial summary

Negative

Negative

 

Negative

 

Positive control

330 ± 31.5

657 ± 18.8

 

269 ± 5.2

 

Strain TA98

 

 

 

 

 

0

19 ± 1.5

26 ± 5.3

54 ± 2.2

26 ± 6.1

 

1

 

 

50 ± 5.8

 

 

3

15 ± 3.0

25 ± 0.7

65 ± 2.0

23 ± 2.7

 

33

18 ± 0.7

58 ± 3.8

70 ± 2.7

22 ± 1.3

 

100

21 ± 1.7

86 ± 12.4

100 ± 19.8

33 ± 2.6

 

333

16 ± 1.9

97 ± 19.9

 

38 ± 0.3

 

Trial summary

Negative

Positive

Positive

Negative

 

Positive control

793 ± 43.1

1,884 ± 71.5

395 ± 3.6

697 ± 40.5

 

*: Revertants are presented as mean ± standard error from three plates.

-Table 2: Mutagenicity of 1,2,3-Trichloropropane in Salmonella typhimurium performed at Microbiological Associate

 

Revertants/plate *

Dose µg/plate

-S9

+10% hamster S9

+10% rat S9

 

Trial 1

Trial 2

Trial 1

Trial 2

Trial 3

Trial 1

Trial 2

0

78 ± 6.5

106 ± 4.7

241 ± 21.1

81 ± 1.9

89 ± 3.8

93 ± 2.3

219 ± 1.0

10

88 ± 1.2

 

527 ± 14.5

762 ± 29.7

728 ± 32.7

176 ± 3.0

 

33

94 ± 2.5

121 ± 2.5

1,008 ± 18.8

1,263 ± 20.0

1,122 ± 29.0

349 ± 10.2

380 ± 7.8

100

86 ± 7.1

106 ± 4.2

1,628 ± 57.7

2,612 ± 269.1

2,728 ± 44.2

748 ± 27.5

700 ± 53.8

333

87 ± 3.8

108 ± 2.7

2,292 ± 136.9

2,879 ± 87.3$

3,235 ± 210.9

1,518 ± 32.7

1,242 ± 54.3

666

115 ± 4.0

 

Toxic

Toxic

148 ± 18.6$

1,924 ± 55.3

 

667

 

121 ± 4.5$

 

 

 

 

1,786 ± 24.2$

1,000

 

Toxic

 

 

 

 

Toxic

Trial summary

Equivocal

Negative

Positive

Positive

Positive

Positive

Positive

Positive control

446 ± 27.0

410 ± 7.2

524 ± 17.9

355 ± 12.7

2,400 ± 65.0

509 ± 17.4

915 ± 26.9

 

 

 

 

 

 

 

 

TA1535

 

 

 

 

 

 

 

0

19 ± 2.4

21 ± 1.7

4 ± 0.6

8 ± 1.9

 

22 ± 2.3

47 ± 4.8

10

14 ± 0.9

 

178 ± 6.7

159 ± 16.5

 

33 ± 1.9

 

33

17 ± 1.9

29 ± 5.5

364 ± 12.3

325 ± 5.9

 

107 ± 2.0

94 ± 4.7

100

19 ± 3.2

24 ± 2.8

786 ± 32.8

720 ± 33.5

 

203 ± 7.9

203 ± 11.5

333

20 ± 3.8

31 ± 2.3

1,286 ± 22.0$

1,340 ± 29.7

 

456 ± 22.6

415 ± 4.2

666

22 ± 2.2

 

Toxic

Toxic

 

549 ± 38.7

 

667

 

23 ± 1.2$

 

 

 

 

544 ± 37.9$

1,000

 

12 ± 0.5$

 

 

 

 

147 ± 20.4$

Trial summary

Negative

Negative

Positive

Positive

 

Positive

Positive

Positive control

280 ± 18.0

330 ± 18.8

59 ± 4.2

256 ± 8.7

 

239 ± 15.2

254 ± 11.9

TA97

 

 

 

 

 

 

 

0

74 ± 2.8

142 ± 4.4

108 ± 6.0

137 ± 3.0

 

111 ± 5.8

183 ± 20.5

10

84 ± 4.2

 

211 ± 6.4

194 ± 6.5

 

133 ± 9.1

 

33

64 ± 8.5

177 ± 2.8

365 ± 5.0

319 ± 20.3

 

162 ± 8.4

233 ± 5.8

100

78 ± 3.5

131 ± 12.2

779 ± 20.1

691 ± 24.7

 

219 ± 12.6

270 ± 7.2

333

93 ± 2.5

160 ± 17.0

1,422 ± 50.3

358 ± 54.5$

 

408 ± 34.4

391 ± 7.3

666

75 ± 2.6

 

270 ± 11.3$

Toxic

 

489 ± 5.0

 

667

 

99 ± 3.8$

 

 

 

 

520 ± 15.2

1,000

 

97 ± 2.0$

 

 

 

 

518 ± 16.1

Trial summary

Negative

Negative

Positive

Positive

 

Positive

Positive

Positive control

105 ± 5.2

345 ± 10.0

521 ± 4.5

532 ± 10.6

 

1,411 ± 29.8

1,307 ± 28.3

TA98

 

 

 

 

 

 

 

0

18 ± 4.6

22 ± 2.4

38 ± 1.2

59 ± 4.9

 

36 ± 6.2

38 ± 1.3

10

19 ± 2.6

 

35 ± 0.3

59 ± 1.5

 

28 ± 3.5

 

33

17 ± 0.6

19 ± 1.9

53 ± 9.6

77 ± 12.5

 

34 ± 0.9

34 ± 2.3

100

18 ± 3.8

24 ± 2.2

76 ± 5.1

82 ± 9.9

 

47 ± 5.2

59 ± 3.0

333

13 ± 2.4

18 ± 2.0

193 ± 7.5

191 ± 24.7

 

67 ± 3.2

68 ± 6.3

666

14 ± 1.8

 

61 ± 8.7$

 

 

89 ± 10.9

 

667

 

22 ± 0.7

 

181 ± 8.7

 

 

91 ± 1.2

1,000

 

Toxic

 

 

 

 

43 ± 3.1$

Trial summary

Negative

Negative

Positive

Positive

 

Positive

Positive

Positive control

189 ± 10.7

219 ± 11.5

2,226 ± 101.1

151 ± 11.0

 

263 ± 11.6

229 ± 11.3

*: Revertants are presented as mean ± standard error from three plates.

$: Slight toxicity

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative without metabolic activation
positive with metabolic activation

1,2,3-trichloropropane was tested in a Bacterial Reverse Mutation Assay (Ames Test) according to OECD Guideline 471 as at 1982, using strains S. typhimurium TA 1535, TA 1537, TA 97, TA 98 and TA 100 in the preincubation assay with and without activation (S9 of hamster and rat) in two different Laboratories.
Test material was mutagenic in S. typhimurium TA 97, TA 98, TA100 and TA1535 with activation at levels >= 1 - 10 µg/plate. It was not mutagenic in any tester strain without S-9 activation, or in TA1537 with S-9. With hamster S9 the sensitivity was usually a bit higher.
Executive summary:

In the present study 1,2,3-trichloropropane at a purity of 99.1 % was tested in a Bacterial Reverse Mutation Assay (Ames Test) according to OECD Guideline 471 as at 1982, using strains S. typhimurium TA 1535, TA 1537, TA 97, TA 98 and TA 100 in the preincubation assay with and without activation (S9 of hamster and rat) in two different Laboratories (Microbial Genetics Department, SRI International 333 Ravenswood Avenue Menlo Park, CA 94025-3493 and Microbiological Associates, 37428, Hills Tech Drive Farmington Hills MI 48331).

Test concentrations were 0, 1, 3, 10, 33, 100, 333 µg/plate (SRI, International) and 0, 10, 33, 100, 333, 666, 1'000 µ g/plate (Microbial Associates). The preincubation assay was conducted using tightly capped incubation vials to account for the volatility of the substance.

The test material was mutagenic in S. typhimurium TA 97, TA 98, TA100 and TA1535 with activation at levels >= 1 - 10 µg/plate. It was not mutagenic in any tester strain without S-9 activation, or in TA1537 with S-9. With hamster S9 the sensitivity was usually a bit higher. Cytotoxicity occured usually at 666 µg/plate.

Based on the obtained results the test material is mutagenic in S. typhimurium TA 98, TA100 and TA1535 with activation at levels >= 1 - 10 µg/plate. It was not mutagenic in any tester strain without S-9 activation, or in TA1537 and TA1538 with S-9. Cytotoxicity to TA 1535 and TA 100 was determined at >= 666 µg/plate.