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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
Study period:
1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Not GLP, purity of the test material was not listed.
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
publication
Title:
Mutagenic evaluation of nitroparaffins in the Salmonella typhimurium/mammalian-microsome test and the micronucleus test
Author:
Hite M and Skeggs H.
Year:
1979
Bibliographic source:
Environ Mutagen 1:383-389

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
The report includes the results of in vitro studies using the method described by Ames (Ames BN, Methods for detecting carcinogens and mutagens with the Salmonella microsome mutagenicity test, Mutat Res 31:347-364, 1975) with and without a rat-liver microsomal activation system.
GLP compliance:
no
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
No data

Method

Target gene:
TA-92- hisG46/pKM101, TA98-hisD3052, TA100- hisG46, TA1537-hisC3076
Species / strain
Species / strain:
other: S. typhimurium strains TA92, TA98, TA100 and TA1537
Metabolic activation:
with and without
Metabolic activation system:
S-9 from Aroclor-treated rats and sodium pentobarbital-treated rats
Test concentrations with justification for top dose:
nominal- 0.0037, 0.011, 0.03, and 0.1 ml per plate (98 mg/plate)

Vehicle:
2-Nitropropane was used as supplied or was diluted with dimethylsulfoxide (DMSO).
Controls
Negative controls:
yes
Solvent controls:
yes
Remarks:
diluent controls
Positive controls:
yes
Positive control substance:
other: 1-methyl-2)3a,4,5,6,7,7a-hexahydro- 1,2-benizisoxazol-3-yl)-5 nitroimidazole (at 7.5 or 150 mg/plate).
Details on test system and conditions:
Bacteria: The tester strains (TA92, TA98, TA100 and TA1537) were stored frozen at -80 degrees C until use. They were thawed and grown for 16 hours at 37 degrees C in a nutrient broth containing 0.5% NaCl.

Test materials: 2-Nitropropane (purity not specified) was used as supplied or was diluted with dimethylsulfoxide (DMSO). The positive control for all tests was 1-methyl-2)3a,4,5,6,7,7a-hexahydro- 1,2-benizisoxazol-3-yl)-5 nitroimidazole (at 7.5 or 150 mg/plate). Concentrations of test material used in strains TA98 and TA100 were 0.0037, 0.11, 0.03 and 0.1 ml/plate (two replicates). All controls and test material concentrations (except for 0.1 ml) were tested in TA98 and TA100 on two separate occasions. The materials were tested once in the other strains. The 0.0037 concentration was not tested in strains TA1537 and TA92.

S-9: The studies were performed with two rat liver microsomal activation preparations. One was derived from CD male rats pretreated i.p. with 75 mg/kg/day sodium pentobarbital for 4 days and fasted for 18 hours. The other preparation was derived from male rats injected i.p. with 500 mg/kg Aroclor 1254 five days before liver isolation. The final incubation mixture contained 0.3 ml of the S-9 supernatant, 8 micromolar MgCl2, 22 micromolar KCl, 5 micromolar glucose-6-phosphate, 4 micromolar triphosphopyridine nucleotide (TPN) and 100 micromolar sodium phosphate (pH 7.4). In the negative control mixtures, S-9 and TPN were replaced with a saline buffer solution.

Test material, DMSO or positive control (0.1 ml), 0.5 ml S-9 mix or buffer and 2 ml of 0.7% seed agar containing 0.5 ml of inoculum/ml were added to test tubes. The cell titer of the inoculum was approximately 10E8 organisms/ml. The test tubes were rotated between the palms and the contents poured onto base plates contining either hisitidine-deficient (N=3) or histidine-supplemented (N=1) medium.

All plates were incubated for 72 hours at 37 degrees C. Growth on the supplemented medium was compared with that of control plates for evidence of bacterial inhibition. Revertants on histidine-deficient plates were enumerated, counts were averages, and were compared with controls.
Evaluation criteria:
The test was considered positive if there was at least a 2-fold increase in revertants that was dose-dependent.

Results and discussion

Test results
Species / strain:
other: S. typhimurium strains TA92, TA98, TA100 and TA1537
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity:
yes
Remarks:
0.1 ml/plate (without S-9), > 0.1 ml/plate (with S-9)
Negative controls valid:
yes
Positive controls valid:
other: Yes, The positive control elicited a positive response at both concentrations in all strains except for TA92 (in the absence or presence of either S-9 preparation) or in TA15347 without S-9.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
All concentrations tested were positive in strain TA100 with or without either S-9 preparation. All concentrations except 0.0037 ml/plate were positive in strain TA98 with or without either S-9 preparation (with the exception of 0.011 ml/plate with S-9 from Aroclor-treated rats and 0.1 ml/plate undiluted material without activation, which was toxic). Undiluted test material (0.1 ml) was positive in TA1537 with either S-9 preparation and was toxic in the absence of S-9. Undiluted test material (0.1 ml) was positive in TA92 with or without either S-9 preparation and 0.03 ml/plate was positive in TA92 in the presence (but not the absence) of either S-9 preparation.

Cytotoxic concentration- 0.1 ml/plate (without S-9), > 0.1 ml/plate (with S-9)


The positive control elicited a positive response at both concentrations in all strains except for TA92 (in the absence or presence of either S-9 preparation) or in TA15347 without S-9.

Number of Revertant Colonies- TA 1537
Dose Aroclor 1254 Phenobarb. No activation
0.1 NP 37.0 33.7 Inhibited
0.03 NP 17.7 21.3 14.7
0.011 NP 14.3 17.3 8.7
150.0 po 113.7 89.3 17.3
7.5 pos 45.3 39.0 12.7
Neg contro 9.1 10.9 7.7

Number of Revertant Colonies- TA 92
Dose Aroclor 1254 Phenobarb. No activation
0.1 NP 215.0 204.3 105.0
0.03 NP 139.0 127.3 53.0
0.011 NP 84.7 83.7 49.0
150 pos 67.7 88.3 38.3
7.5 pos 58.0 51.0 42.7
Neg control 46.8 56.2 33.5

Number of Revertant Colonies- TA 98
Dose Aroclor 1254 Phenobarb. No activation
0.1 NP 252.0 246.7 Inhibited
0.03 NP 204.3, 195.3 198.3, 185.7 102.0, 133.0
0.011 NP 131.0, 95.0 127.3, 156.0 75.7, 74.0
0.0037 NP 96.0 106.3 49.3
150 pos 573.3, 753.7 505.3, 469.3 110.0, 98.0
7.5 pos 413.3, 234.3 238.3, 236.3 63.7, 56.3
Neg control 57.6, 54.4 54.9, 64.1 28.2, 32.9

Number of Revertant Colonies- TA 100
Dose Aroclor 1254 Phenobarb. No activation
0.1 NP 501.3 397.3 Inhibited
0.03 NP 431.0, 547.0 366.3, 407.7 223.7, 421.7
0.011 NP 275.7, 327.3 242.0, 312.7 160.7, 278.7
0.0037 NP 193.0 198.0 143.3
150 pos 1276.0, 1216.0 1336.0, 1289.7 917.3, 1070.3
7.5 pos 1484.0, 1425.7 1330.7, 1247.0 323.7, 436.0
Neg control 66.1, 68.5 60.6, 71.2 76.1, 81.6

Any other information on results incl. tables

Triphosphopyridine nucleotide is not usually present in S-9.  It is unknown if this had an effect on the results.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive

2-Nitropropane is positive in the Ames assay.
Executive summary:

None