Semicarbazide hydrochloride

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

No GLP.

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1983

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

1. Four bacteria strain were used in the study. According the TG 471 - 5 strains of bacteria should be used.

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Semicarbazide hydrochloride

EC Number:

209-247-0

EC Name:

Semicarbazide hydrochloride

Cas Number:

563-41-7

Molecular formula:

CH5N3O.ClH

IUPAC Name:

semicarbazide hydrochloride

Method

Target gene:

Histidine-requiring gene in Salmonella typhimurium.

Species / strain

Species / strain / cell type:

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

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9 rat and hamster liver

Test concentrations with justification for top dose:

At least five doses were used in each strain in the presence of absence S9.
Test substance 33.0, 100, 333, 1000, 2167 ug/plate.
Positive control 1.5 ug/plate in case of rats S9 activation and 0.75 ug/plate in case of hamster S9 activation.

The highest dose was selected according the preliminary study,where chemicals were checked for toxicity to TAl00 up to a concentration of 10 mg/plate or the limit of solubility, both in absence and in presence of S-9mix. As the indicator of toxicity was used Viability of complete medium and reduced numbers of revertant colonies per plate and/or thinning or absence of the bacterial lawn. If toxicity was not apparent in the preliminary toxicity determination, the highest dose tested was 10 mg/plate; otherwise the upper limit of solubility was used. If toxicity was observed, the doses of test chemical were chosen so that the high dose exhibited some degree of toxicity. Occasionally, in the earlier tests, the high dose was greater than 10 mg/plate.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: test substance is compatible with the survival of the bacteria and the S9 activity.

Controls

Untreated negative controls:

yes

Remarks:

choline chloride, glycerol, glycine, mannitol, and sodium phosphate

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Positive control substance:

9-aminoacridine

sodium azide

other: 2 Amino antracene tested on all strains

Details on test system and experimental conditions:

METHOD OF APPLICATION:
Preincubation Methodology
Briefly, 0.5 ml of S-9 mix or 0. 1 M PO4 buffer was dispensed into an appropriate number of 13 X 100 mm culture tubes maintained at 37°C in a dry-bath. Then, 0.05 ml of cells and 0.05 ml of solvent or chemical dilution were added to each tube. The mixture was vortexed and allowed to incubate with shaking in the early tests (CWR, EGG), or standing (SRl) for 20 min at 37°C. The protocol was later changed to eliminate the shaking procedure, because the commercial shakers available would not fit in the Class 11. Type B hoods and, for the purposes of laboratory safety, it was inadvisable to incubate the chemicals at 37°C in the open laboratory. Following the preincubation period, 2.5 ml (EGG) or 2.0 ml (CWR, SRI) of molten top agar (45°C) supplemented with 0.5 mM L-histidine and 0.5 mM d-biotin was pipetted into the tubes, which were immediately vortexed, and their contents poured onto 25 ml of minimal glucose bottom agar in a 15 x 100-mm plastic petri dish (Falcon Muta-Assay, 1028 [EGG, SRI] or Fisher Scientific petri dishes [CWR]). After the overlay solidified, the plates were inverted and incubated at 37°C for 48 h.

DURATION
- Preincubation period:Yes,20 minutes at 37ºC
- Exposure duration: 48 hour at 37ºC

NUMBER OF REPLICATIONS: Three plates were used, and the experiment was repeated no less than 1 week after completion of the initial test.

Statistics:

If a chemical was mutagenic or gave a questionable response, it was analyzed by Radian for identity and purity. Analyses had been performed previously by Midwest Research Institute (MRI) on selected other chemicals and on chemicals that had been tested in the National Cancer Institute’s Carcinogenesis Bioassay Program

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:

not specified

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:

not specified

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:

not specified

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:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

Test item do not induce any mutagenic change in tested bacterias (Salmonella typhimurium TA98, TA100, TA 1535 and TA 1537) with and without metabolic activation.

Executive summary:

According the present study test substance was dissolved in water and tested with the pre-incubation procedure with S.typhimurium strains TA1535, TA1537, TA98,TA100 with and without metabolic activation S9. To select the dose range, chemicals were checked for the toxicity to TA100 strain in presence and in absence of S9 obtained from rat and mouse liver.All chemcials were tested used preincubation period at the temperature 37ºC in a dry bath for 20 minutes. After the overlay solidified, the plates were inverted and incubated at 37°C for 48 h. Positive and negative solvent controls were used also. There was no evidence of any increase in the number of revertant colonies in the presence of the test item solution and dilutions without and with metabolic activation. Based on the result of this study, the test item was not found to be mutagenic under the test conditions.