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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:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
The restrictions were that only a short exposure time was used. It was not compliant with GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1978
Report date:
1978

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
yes
Remarks:
only short exposure time used
GLP compliance:
no
Type of assay:
mammalian cell gene mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Potassium salts of [hexane-1,6-diylbis[nitrilobis(methylene)]]tetrakisphosphonic acid (4-7 K:1)
EC Number:
701-184-1
Molecular formula:
HMDTMP-4K C10H24K4N2O12P4 HMDTMP-5K C10H23K5N2O12P4 HMDTMP-6K C10H22K6N2O12P4 HMDTMP-7K C10H21K7N2O12P4
IUPAC Name:
Potassium salts of [hexane-1,6-diylbis[nitrilobis(methylene)]]tetrakisphosphonic acid (4-7 K:1)
Test material form:
liquid

Method

Target gene:
Thymidine kinase
Species / strain
Species / strain / cell type:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Metabolic activation system:
Aroclor induced rat liver S9
Test concentrations with justification for top dose:
5-30 µl/ml
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: none given in study report
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
without metabolic activation 0.5 µl/ml
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: dimethylnitrosamine 0.3 µl/ml
Remarks:
with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 4 hours
- Expression time (cells in growth medium): 3 days
- Selection time (if incubation with a selection agent): 10 days

SELECTION AGENT (mutation assays): selection medium containing 5% v/v BrdU

NUMBER OF REPLICATIONS: triplicate plates

DETERMINATION OF CYTOTOXICITY
- Method: cloning efficiency; relative total growth

ACTIVATION: NADP and isocitric acid were used as co-factors. Final concentration of S9 is not clearly indicated in the report.
Evaluation criteria:
A substance is considered mutagenic if there is a dose-related increase over 3 concentrations with the minimum increase at least 2.5 times the solvent and/or negative control values.

Results and discussion

Test results
Key result
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
20 µl/ml
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

Results of mutagenicity assay (mean of three plates)

Concentration µl/ml

Relative cloning efficiency

Relative total growth

Mutant frequency

-MA

+MA

-MA

+MA

-MA

+MA

Negative control

100

100

100

100

9.0

7.0

Solvent control

92.2

102.4

67.0

102.8

15.5

14.3

5

68.4

71.7

74.7

81.2

8.8

8.6

10

79.9

100.8

62.2

80.8

6.4

14.1

15

73.6

82.5

64.4

83.5

15.6

7.3

20

60.0

40.7

57.1

36.7

13.6

4.3

30

5.4

2.0

3.9

2.2

28.3

9.6

Positive control

35.4

7.3

18.0

0.3

180.4

700

Applicant's summary and conclusion

Conclusions:
HMDTMP (4-7K) has been tested according to a protocol that is similar to OECD 476. No increase in the mutant frequency was observed at any concentration up to cytotoxic concentrations, with and without metabolic activation. Appropriate positive, solvent and negative controls were included and gave the expected results. It is concluded that the substance is negative for mutagenicity to L5178Y mouse lymphoma cells under the conditions of the test.