N-hydroxysuccinimide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1986

Report date:

1985

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

only four strains tested

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

N-hydroxysuccinimide

EC Number:

228-001-3

EC Name:

N-hydroxysuccinimide

Cas Number:

6066-82-6

Molecular formula:

C4H5NO3

IUPAC Name:

1-hydroxypyrrolidine-2,5-dione

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source: Chemical Dynamics (South Plainfield, NJ).

Method

Target gene:

Histidine

Species / strain

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

Bacterial strains were received from B.N. Ames.

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

0, 100, 500, 1000 and 5000 µg/plate.
No toxicity was observed at the highest dose (5000 µg/plate).

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: the test item is completely soluble in the vehicle

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

other: N-methyl-N'-nitro-N-nitrosoguanidine; 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)
- Cell density at seeding (if applicable): 1E08 cells/ml
The plate incorporation assay was conducted accoding to Ames et al. (1975). To 2 ml of molten top agar at 45ºC were added 0.1 ml bacterial culture containing approximately 1E08 cells/ml, 0.1 ml test solution or solvent and 0.5 ml S) mix or buffer. After mixing, the soft agar was poured onto selective minimal agar plates.

DURATION
- Exposure duration: 2 days at 37 ºC

SELECTION AGENT (mutation assays): lack of histidine in the media

NUMBER OF REPLICATIONS: 2

DETERMINATION OF CYTOTOXICITY
- Method: Toxicity was stimated by examining background bacterial lawns using a stereoscope.

- OTHER:
Preparation of the metabolic activation system: S9 was prepared from aroclor 1254-induced male Sprague-Dawley rat livers.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535

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

Key result

Species / strain:

S. typhimurium TA 1537

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 3

Key result

Species / strain:

S. typhimurium TA 98

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 4

Key result

Species / strain:

S. typhimurium 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

Any other information on results incl. tables

Table 1.Number of revertant colonies after exposure to N-hydroxisuccinimide or positive control with metabolic activation.

 

Strain	Positive control	Amount tested (µg/plate)	Mean number of colonies per plate
			N-hydroxysuccinimide	Positive control
TA 1535	-	5000	11	-
	-	1000	11	-
	-	500	14	-
	-	100	10	-
	-	0	13	-
	2-AA	10	-	183
TA 1537	-	5000	9	-
	-	1000	5	-
	-	500	7	-
	-	100	9	-
	-	0	11	-
	2-AA	50	-	175
TA 98	-	5000	30	-
	-	1000	28	-
	-	500	26	-
	-	100	28	-
	-	0	35	-
	2-AA	20	-	1137
TA 100	-	5000	168	-
	-	1000	184	-
	-	500	176	-
	-	100	176	-
	-	0	211	-
	2-AA	10	-	1535

2-AA: 2-aminoanthracene

 

Table 2.Number of revertant colonies after exposure to N-hydroxisuccinimide or positive control without metabolic activation.

 

Strain	Positive control	Amount tested (µg/plate)	Mean number of colonies per plate
			N-hydroxysuccinimide	Positive control
TA 1535	-	5000	15	-
	-	1000	10	-
	-	500	15	-
	-	100	12	-
	-	0	13	-
	MNNG	5	-	1190
TA 1537	-	5000	4	-
	-	1000	7	-
	-	500	7	-
	-	100	5	-
	-	0	7	-
	9-AA	5	-	322
TA 98	-	5000	19	-
	-	1000	22	-
	-	500	15	-
	-	100	23	-
	-	0	15	-
	2-NF	5	-	959
TA 100	-	5000	160	-
	-	1000	163	-
	-	500	157	-
	-	100	174	-
	-	0	172	-
	MNNG	5	-	1055

MNNG: N-methyl-N’-nitro-N-nitrosoguanidine

9-AA: 9-aminoacrinide

2-NF: 2-nitrofluorene

Applicant's summary and conclusion

Conclusions:

The test item did not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98 and TA 100 with and without metabolic activation up to 5000 µg/plate.

Executive summary:

A study to determine the ability of the test item to induce mutation was assessed by the bacterial reverse mutation test, performed according to the method described by Ames, similar to OECD 471. Four histidine dependent strains of Salmonella typhimurium (TA 1535, TA1537, TA 98 and TA 100) were exposed to 0, 100, 500, 1000 and 5000 µg/plate of the test item in presence and absence of S9 rat liver metabolic activation, by the plate incorporation method. Under the experimental conditions used, the test item did no induce an increase in the number of revertants in any strain at any dose level, either with or without metabolic activation. Therefore, the test item was not mutagenic.