Semicarbazide hydrochloride

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Weight of evidence: Ames test: 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.

In another study, test substance showed negative results with respect to genotoxicity in the DNA repair test with hepatocytes for both species (rat and mice), negative results in mutagenicity in Salmonella Typhymurrium but positive results in carcinogenicty.

Link to relevant study records

Reference

Reference 1

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.

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

Target gene:

Histidine-requiring gene in Salmonella typhimurium.

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.

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

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

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

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

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

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.

Reference 2

Endpoint:

in vitro DNA damage and/or repair study

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

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 482 (Genetic Toxicology: DNA Damage and Repair, Unscheduled DNA Synthesis in Mammalian Cells In Vitro)

Deviations:

no

GLP compliance:

no

Type of assay:

other: Hepatocyte/DNA repair test in mammalian cells in vitro

Species / strain / cell type:

hepatocytes: Rat and mouse

Details on mammalian cell type (if applicable):

CELLS USED
- Source of cells:isolated from the liver of rats (ACI/N) weighting 200 -250 g and from liver of mice (C3H/HeN) weighing 20 g

MEDIA USED
- Type and identity of media including CO2 concentration if applicable:William´s medium E

Additional strain / cell type characteristics:

not specified

Test concentrations with justification for top dose:

0.001,0.0001,0.00001,0.000001 and 0.0000001 M.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: water

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

9,10-dimethylbenzanthracene

other: N-2fluorenylacetoamide

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium-Williams Medium E.

DURATION
- Exposure duration:20 hours
- Selection time (if incubation with a selection agent):14 days

NUMBER OF REPLICATIONS:2

NUMBER OF CELLS EVALUATED: 50 cells/ coverslips

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

Evaluation criteria:

Each cell nucleus was positive when the net nuclear grain count was more than 5 grains above the cytopasmic background.
Test substance was considered positive when the mean nuclear grain count was more than 5 grains or positive cells were more tna 33%.

Statistics:

Autoradiographic grains were counted on a CRT display (Olympus type S) with microscopic attachment.

Key result

Species / strain:

hepatocytes: Rat or mice

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Table of results:

Chemical	Dose (M)	Rat		Mouse
		UDS grains/nucleus	DNA repair	UDS grains/nucleus	DNA repair
Semicarbazide hydrochloride	10^-3	-1.2+-1.6	-	-0.6+-1.3	-
	10^-4	-0.4+-1.5		-0.7+-1.2
	10^-5	-1.2+-1.7		-0.8+-1.2

- : negative

Positive DNA response were not present at two doses (10 -6 and 10 -7M) they are not shown in the table.

Conclusions:

Test substance was negative with respect to genotoxicity in the DNA repair test with hepatocytes for both species (rat and mice) and also it had negative results in mutagenicity in Salmonella typhimurium strains.

Executive summary:

In the present study, semicarbazide hydrochloride was tested to genotoxicity in the DNA repair test with hepatocytes from rats and mices. As medium was used Williams Medium E. After hepatocytes were isolated, they were attached to plastic coverslips in primary culture for 2 hours using Williams Medium E. Then were washed and exposed to the test compunds for 20 hours. Test item was dilluted with distilled water before addition to the cultures (10-3 and 10-7 M). Concentration tested included the highest soluble non toxic dose. At the end of incubation, the cultures were washed in medium and the coverslips were mounted on glass sides which were dipped in Sakura NR-M2 photographic emulsions and exposed for 14 days. In the concentration of 10-6 and 10 -7 DNA reponse was not presented. Based on the results obtained from that study, test substance is negative with respect to genotoxicity in the DNA repair test with hepatocytes for both species (rat and mice).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Weight of evidence: Substance tested in vivo micronucleus assay does not show any visual signs of distress (fur condition, wakefulness, etc) nor adverse effects in two different mouse strains.

In another vivo study (UDS assay), test item does not induce mutagenicity to liver UDS of female CD-1 mice in a single oral dose of 100 or 200 mg/kg bw.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

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

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

Five animals per each group should be used. In the study, 3 and 4 animals were used per each group (Balb/C mice)

GLP compliance:

no

Type of assay:

mammalian germ cell cytogenetic assay

Species:

mouse

Strain:

other: CBA and Balb/C

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Scanbur BK, Sollentuna, Sweden
- Age at study initiation: CBA - 6-8 weeks old, Balb/C - 6-7 weeks ols
- Weight at study initiation: CBA - 20-25 g, Balb/C - 15-25 g
- Assigned to test groups randomly: Yes,randomly divided into the different dose groups.
- Diet (e.g. ad libitum): Yes,standard diet , free access
- Water (e.g. ad libitum):Yes, tap water, free access

Route of administration:

intraperitoneal

Details on exposure:

Semicarbazide and positive control (Colchicine) were disolved in PBS to give solutions with different cocnentrations of the tested compound. All mice were given an acute single i.p. (10 ul/g b.w.) injection with test substance, negative control and positive control.

Post exposure period:

Blood samples were collected at 42 hours after injections.

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Experiment 1

Dose / conc.:

40 mg/kg bw/day (nominal)

Remarks:

Experiment 1

Dose / conc.:

80 mg/kg bw/day (nominal)

Remarks:

Experiment 1

Dose / conc.:

120 mg/kg bw/day (nominal)

Remarks:

Experiment 1

Dose / conc.:

80 mg/kg bw/day (nominal)

Remarks:

Experiment 2

Dose / conc.:

120 mg/kg bw/day (nominal)

Remarks:

Experiment 2

No. of animals per sex per dose:

Experiment 1 - each dose group - Three of four animals.
Experiment 2 - 5 per group.

Control animals:

yes, concurrent no treatment

Positive control(s):

Colchicine
- Route of administration: INTRAPERITONEAL
- Doses / concentrations: 1.0 mg/kg b.w

Tissues and cell types examined:

Peripheral blood of mice.

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: Limited pre-study was carried out to set the maximum tolerated dose.

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Blood samples were collected, under light anaesthesia with Fluothane (Zeneca, Goteborg), from the orbital plexus of each animal, both CBA and Balb/C mice, at 42 h after injections. The choice of the sampling time is based on the knowledge that the time between the appearance of polychromatic erythrocytes (PCE) in bone marrow and peripheral blood is about 20 h. From each animal, about 50 ul of blood was drawn. Three parallel aliquots of 5ul of blood were layered for purification on a 65% Percoll (Pharmacia Biosystems, Uppsala, Sweden) gradient.

METHOD OF ANALYSIS:
Flow cytometry: Samples were analyzed on FACSVantage SE flow cytometer (BD Immunocytometry systems, Sunnyvale, CA) equipped with an argon ion laser (Enterprise II, Coherent, Santa Clara, CA) operating at both multiline UV and 488 nm. UV output power was 50 mW. Cells, one by one, pass the two beams. Here, the analysis rate was about 1000 cells per second. From each cell different electric signals are sent out, e.g. from forward scatter (FSC) and side scatter (SSC) information’s about structure and size are given. In this analysis a threshold was set in FSC to include all intact cells. Using CellQuest software, peak values for FSC, SSC, Thiazole orange fluorescence and Hoechst 33342 (HO342) fluorescence signals were collected. The FSC signals were acquired using a linear scale and the SSC, TO and HO342 signals were acquired using a log scale.
From each sample between 20 000 and 100 000 events (cells) were collected. CellQuest software (BD) was used for data acquisition and analysis.

For the calculation of MPCE frequencies, dot plots of FSC versus SSC, and DNA content (HO342), indicating the presence of MN, versus RNA (TO) content, indicating the age of the erythrocytes were displayed for each analysed sample.

The cells in the pellet were fixed in a solution of glutaraldehyde. The fixed cells were then stored during 2–5 days at 4 ◦C and the following staining was made in a buffer prepared by adding the fluorescent dyes Hoechst 33342 (HO 342) (Sigma–Aldrich, Sweden) and Thiazole orange (TO) (Molecular probes, Eugene, OR, USA) to PBS. HO
342 is a DNA dye and TO is a RNA dye.

Statistics:

The two-tailed, Student t-test was used for a statistical evaluation of the results.

Key result

Sex:

male

Genotoxicity:

negative

Toxicity:

not specified

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose range: 200 (four mice), 80 (three mice), and 50 mg/kg b.w. (three mice).
- Clinical signs of toxicity in test animals: The highest dose (200 mg/kg b.w.) of semicarbazide hydrochloride - all four tested mice died within 1 hour. At the doses of 80 mg/kg or 50 mg/kg b.w. - no sign of adverse effect was noticed.
On the bases of these results and LD50 from another study, the selected dose in the micronucleus test were 40,80 and 120 mg/kg bw.

Results from the micronucleus assay with semicarbazide in Experiment 1

Strain	Animal code	Dose Mg/kg bw	%PCE	S.D.	PCE	Number of MPCE	fMPCE per mille	S.D.
Balb/C	101	0	20		204 589	400	1.95
Balb/C	102	0	2.3		220 700	345	1.56
Balb/C	103	0	3.6		224 589	434	1.93
Balb/C	104	0	2.3		221 667	392	1.77
Balb/C	101-104	0	2.5	0.72	871 545	1571	1.8	0.18
Balb/C	105	40	1.5		96 745	166	1.71
Balb/C	106	40	1.7		209 819	355	1.69
Balb/C	107	40	2.3		216 084	399	1.84
Balb/C	105-107	40	1.8	0.40	522 648	920	1.76	0.08
Balb/C	112	80	2.3		220 071	380	1.72
Balb/C	113	80	1.7		201 970	381	1.88
Balb/C	114	80	2.1		202 810	434	2.14
Balb/C	112-114	180	2.1	0.33	624 851	1195	1.91	0.34
Balb/C	115	120	2.2		222 684	413	1.85
Balb/C	116	120	2.2		208 059	422	2.02
Balb/C	117	120	2.4		226 696	502	2.21
Balb/C	118	120	2.1		216 086	435	2.01
Balb/C	115-118	120	2.2	0.17	873 525	1772	2.02	0.12
Balb/C	118	Colch.	0.8		30 217	216	7.10
Balb/C	109	Colch.	1.0		58 302	376	6.41
Balb/C	110	Colch.	0.9		44 331	401	8.96
Balb/C	108-110	Colch.	0.8*	0.14	132 850	993	7.4*	1.32

∗ p < 0.001 (Student t-test, two-tailed).

Results from the micronucleus assay with semicarbazide in Experiment 2

Strain	Animal code	Dose Mg/kg bw	%PCE	S.D.	PCE	Number of MPCE	fMPCE per mille	S.D.	Number of MPCE (II)	fMPCE (II) per mille	S.D.
CBA	1	0	2.3		65 645	68	1.03		38	0.58
CBA	2	0	1.6		170 903	237	1.38		112	0.65
CBA	3	0	2.2		201 091	201	1.00		96	0.48
CBA	4	0	2.4		213 483	217	1.02		103	0.48
CBA	5	0	2.0		191 284	166	0.87		76	0.40
CBA	1-5	0	2.0	0.31	842 406	889	1.05	0.9	425	0.50	0.010
CBA	6	80	1.7		176 295	168	0.95		88	0.50
CBA	7	80	2.2		230 744	222	0.96		107	0.46
CBA	8	80	2.1		245 383	242	0.99		119	0.48
CBA	9	80	1.9		224 674	197	0.88		87	0.39
CBA	10	80	1.8		193 270	165	0.85		79	0.41
CBA	6-10	80	1.9	0.21	1 070 366	994	0.93	0.06	480	0.45	0.05
CBA	11	120	2.0		219 505	199	0.91		98	0.45
CBA	12	120	1.6		206 114	171	0.83		77	0.37
CBA	13	120	2.0		204 669	178	0.87		88	0.45
CBA	14	120	1.9		196 744	181	0.92		92	0.47
CBA	15	120	2.3		237 726	186	0.78		78	0.33
CBA	11-15	120	2.0	0.23	1 064 758	915	0.86	0.06	433	0.41	0.06
CBA	16	Colch.	0.46		22 381	88	3.92		62	2.76
CBA	17	Colch.	0.74		34 740	137	3.93		98	2.81
CBA	18	Colch.	0.55		29 939	112	3.73		81	2.70
CBA	1-18	Colch.	0.6*	0.14	87 060	337	3.9*	0.11	241	7.86*	0.06

∗ p < 0.001 (Student t-test, two-tailed).

Conclusions:

The test substance does not show any visual signs of distress (fur condition, wakefulness, etc) nor adverse effects in two different mouse strains.

Executive summary:

According the present study, test substance was tested for genotoxicity in the flow cytometry-based micronucleus assay in vivo. Two strains of male mice were used (CBA and Balb/C). They were injected intraperitonealy with test substance. Before the main test, range finding study was performed to set the maximum tolerated dose. Positive (Colchicine) and negative control (PBS) groups were used too. Test substance and positive control (Colchicine) were disolved in PBS to give solutions with different cocnentrations of the tested compound. All mice were given an acute single i.p. (10 ul/g b.w.) injection with test substance, negative control and positive control.The mice were randomly divided into groups. Two experiments were carried out. Experiment 1 with doses 0, 40,80 and 120 mg/kg/bw and experiment 2, only with the highest doses (80 and 120 mg/kg bw). Blood samples were collected and the frequency of micronucleated polychromatic erythrocytes (MPCE), fMPCE and the cell proliferation, % PCE was determinated. All animals were observed during the experiment for side effects of the treatment. Based on the results from different analyses, the test substance has negative effect on tested animals.

Reference 2

Endpoint:

in vivo mammalian cell study: DNA damage and/or repair

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Remarks:

No GLP, no Guideline followed

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 486 (Unscheduled DNA Synthesis (UDS) Test with Mammalian Liver Cells in vivo)

Deviations:

no

GLP compliance:

no

Type of assay:

unscheduled DNA synthesis

Species:

mouse

Strain:

CD-1

Sex:

female

Route of administration:

oral: gavage

Duration of treatment / exposure:

2 and 16 hours

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

200 mg/kg bw/day (nominal)

Control animals:

yes

Key result

Sex:

female

Genotoxicity:

negative

Toxicity:

not specified

Conclusions:

Test item does not induce mutagenicity to liver UDS of female CD-1 mice in a single oral dose of 100 or 200 mg/kg bw.

Executive summary:

According the data from the mentioned publication, mice were administered with SEM-HCl orally at doses of 100 or 200 mg/kg bw (doses of 67 or 135 mgSEM/kg bw). Liver cells were isolated cultured at 2 and 16 h after dosing and analysed for incorporation of 3H-thymidine. Also, the positives controls were used in that study. Test substance, semicarbazide hydrochloride, showed no effect to female CD-1 mice.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

IN VITRO studies:

Ames test (Haworth et al)

Test substance was dissolved in water and tested with the plate incorporation method with S.typhimurium strains TA1535, TA1537, TA98,TA100 with and without metabolic activation S9. Firstly, chemicals were checked for the toxicity to TA100 strain in presence and in absence of S9 obtained from rat and mouse liver to check the toxicity. Prior to test, the preincubation period at the temperature 37ºC in a dry bath for 20 minutes was used. No mutagenicity was observed in any bacterial strains.

UDS assay (Mori K.):

Test substance was tested to genotoxicity in the DNA repair test with hepatocytes from rats and mices. As medium was used Williams Medium E. After hepatocytes were isolated, they were attached to plastic coverslips in primary culture for 2 hours using Williams Medium E. Then were washed and exposed to the test compunds for 20 hours. Test item was dilluted with distilled water before addition to the cultures (10-3 and 10-7 M). Concentration tested included the highest soluble non toxic dose. Test substance is negative with respect to genotoxicity in the DNA repair test with hepatocytes for both species (rat and mice), negative results in mutagenicity in Salmonella Typhymurrium but positive results in carcinogenicty.

IN VIVO studies:

Micronucleus assay (Zetterberg A.L.)

Test substance was tested for genotoxicity in the flow cytometry-based micronucleus assay in vivo. Two strains of male mice were used (CBA and Balb/C). They were injected intraperitonealy with test substance. Before the main test, range finding study was performed to set the maximum tolerated dose. Positive (Colchicine) and negative control (PBS) groups were used too. Test substance and positive control (Colchicine) were disolved in PBS to give solutions with different cocnentrations of the tested compound. All mice were intraperitoneally injected (10 ul/g b.w.) with test substance, negative control and positive control. Two experiments were carried out. Experiment 1 with doses 0, 40,80 and 120 mg/kg/bw and experiment 2, only with the highest doses (80 and 120 mg/kg bw). Blood samples were collected and the frequency of micronucleated polychromatic erythrocytes (MPCE), fMPCE and the cell proliferation, % PCE was determinated. All animals were observed during the experiment for side effects of the treatment. Based on the results from different analyses, the test substance has negative effect on tested animals.

UDS assay (Nestmann E.R)

According the data from the present study, mice were administered with SEM-HCl orally at doses of 100 or 200mg/kg bw. Liver cells were isolated cultured at 2 and 16 h after dosing and analysed for incorporation of 3H-thymidine. Positives controls were used in that study. Test substance, semicarbazide hydrochloride, showed no effect to female CD-1 mice.

.

Justification for classification or non-classification

Based on the data obtained from several in vitro and in vivo studies semicarbazide hydrochloride is not classified for cell mutagenicity according to CLP Regulation no. 1272/2008.