Disodium 3,3'-dithiobis[propanesulphonate]

Genetic toxicity: in vitro

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:

1 July - 1 August 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EEC Methods for Determination of Toxocity , Annex to Directive 92/69/EEC , Method B14 , Other effects - Mutagenicity : Samonella typhimurium - Reverse Mutation Assay

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: conforms to the guidelines for bacterial mutagenicity testing by the major Japanese Regulatory Authorities including MITI , MOL and MAFF

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

(The Department of Health of the Government of the United Kingdom)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Disodium 3,3'-dithiobis[propanesulphonate]

EC Number:

248-324-3

EC Name:

Disodium 3,3'-dithiobis[propanesulphonate]

Cas Number:

27206-35-5

Molecular formula:

C6H14O6S4.2Na

IUPAC Name:

disodium 3,3'-disulfanediyldipropane-1-sulfonate

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): SPS; 1-Propanesulfonic acid, 3,3´-dithiobis-, disodium salt
- Substance type: white powder
- Storage condition of test material: room temperature

Method

Target gene:

his operon (Salmonella strains)
trp operon (Escherichia coli strain)

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

incubated in the presence of a induced liver microsomal fraction (S9) prepared from rats

Test concentrations with justification for top dose:

First and second experiment : 50 , 150 , 500 , 1500 and 5000 µg/plate with and without metabolic activation

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: sterile distilled water
- Justification for choice of solvent/vehicle: the test material was soluble in water up to 50 mg/ml

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

See -Any other information on materials and methods incl. tables- for details

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium; in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: triplicates

NUMBER OF CELLS EVALUATED: 3 plates per dose level , in two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: Assessment for numbers of revertant colonies and examination for effects on the growth of the bacterial background lawn .


OTHER: In a range-finding study , five concentrations (50,150,500,1500 and 5000 µg/plate) were assayed in triplicate against each tester strain (with and without S9-mix) , using the direct plate incorporation method . Plates were incunated at 37°C for 48 h .

Evaluation criteria:

For a substance to be considered positive in this test system , it should have induced a dose-related and statistically significant increase in the revertant count in one or more strains of bacteria in the presence and/or absence of S9 in both experiments at sub-toxic dose levels .
To be considered negative , the number of revertants at each dose level should be less than twofold to that of the vehicle control frequency .

Statistics:

A statistical analysis of the data was not required to determine the result of the test .

Results and discussion

Test results

Test results 1

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:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES: Results for the negative controls (spontaneous mutation rates) were considered to be acceptable .

Remarks on result:

other: all strains/cell types tested

Any other information on results incl. tables

Table 1 : Range-finding study

		Mean number of revertant colonies per plate (average of 3 plates)
With or without S9 -Mix	Test substance concentration (µg/plate)	Base-pair substitution type			Frameshift type
		TA 100	TA 1535	WP2uvrA^-	TA 98	TA 1537
-	0	103	18	20	24	10
-	50	98	20	18	21	7
-	150	103	17	16	17	8
-	500	104	14	19	22	8
-	1500	95	19	17	23	9
-	5000	105	24	16	23	12
Positive controls , -S9	Name	ENNG	ENNG	ENNG	4NQO	9AA
	Concentrations (µg/plate)	3	5	2	0.2	80
	Mean No. of colonies/plate (average of 3)	490	636	653	206	887
+	0	107	15	23	30	13
+	50	103	14	23	26	12
+	150	101	15	23	29	13
+	500	103	14	24	24	13
+	1500	101	18	21	28	13
+	5000	110	20	23	27	12
Positive controls , +S9	Name	2AA	2AA	2AA	2AA	2AA
	Concentration (µg/plate)	1	2	10	0.5	2
	Mean No. of colonies)plate (average of 3)	1063	241	822	367	236

ENNG = N-ethyl-N´-nitro-N-nitrosoguanidine

4NQO = 4-nitroquinoline-1 -oxide

9AA = 9-aminoacridine

2AA = 2-aminoanthracene

Table 2 : Main study

		Mean number of revertant colonies per plate (average of 3 plates)
With or without S9 -Mix	Test substance concentration (µg/plate)	Base-pair substitution type			Frameshift type
		TA 100	TA 1535	WP2uvrA^-	TA 98	TA 1537
-	0	86	15	17	18	8
-	50	81	14	17	15	7
-	150	88	14	17	19	7
-	500	82	16	17	20	5
-	1500	91	16	17	18	6
-	5000	90	15	19	18	9
Positive controls , -S9	Name	ENNG	ENNG	ENNG	4NQO	9AA
	Concentrations (µg/plate)	3	5	2	0.2	80
	Mean No. of colonies/plate (average of 3)	477	687	958	256	858
+	0	106	16	21	25	11
+	50	103	14	21	30	9
+	150	101	16	24	27	10
+	500	98	16	19	28	7
+	1500	111	17	21	30	8
+	5000	94	15	22	26	8
Positive controls , +S9	Name	2AA	2AA	2AA	2AA	2AA
	Concentration (µg/plate)	1	2	10	0.5	2
	Mean No. of colonies)plate (average of 3)	1126	331	769	422	235

ENNG = N-ethyl-N´-nitro-N-nitrosoguanidine

4NQO = 4-nitroquinoline-1 -oxide

9AA = 9-aminoacridine

2AA = 2-aminoanthracene

Applicant's summary and conclusion

Conclusions:

Interpretation of results:
negative with metabolic activation
negative without metabolic activation

The study was performed according to the OECD TG471 without deviations and therefore considered to be of the highest quality (reliability Klimisch 1). The validity criteria of the test system are fulfilled. No significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains , with any dose of the test material , either with or without metabolic activation . The test material was considered to be non-mutagenic under the conditions of this test.

Executive summary:

Salmonella typhimurium strains TA1535, TA1537, TA98 and TA100 and Escherichia coli strain WP2uvrA- were treated with the test material using the Ames plate incorporation method at five dose levels, in triplicate, both with and without the addition of a rat liver homogenate metabolising system (10% liver S9 in standard co-factors). This method conforms to the guidelines for bacterial mutagenicity testing published by the major Japanese Regulatory Authorities including MITI, MHW, MOL and MAFF. lt also meets the requirements of the OECD, EC and USA, EPA (TSCA) guidelines. The dose range was determined in a preliminary toxicity assay and was 50 to 5000 µg/plate in the first experiment. The experiment was repeated on a separate day using the same dose range as experiment 1, fresh cultures of the bacterial strains and fresh test material formulations. The vehicle (sterile distilled water) control plates gave counts of revertant colonies within the normal range. All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies, both with and without metabolic activation. The test material caused no visible reduction in the growth of the bacterial background lawn at any dose level. The test material was, therefore, tested up to the maximum recommended dose level of 5000 µg/plate. No significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains, with any dose of the test material, either with or without metabolic activation. The test material was considered to be non-mutagenic under the conditions of this test.