Acetamide, N-(2,4-dinitrophenyl)-, reaction products with p-phenylenediamine, sodium sulfide (Na2(Sx)) and sulfur, leuco deriv.

Endpoint summary

Description of key information

In an acute oral toxicity study in rats the acute oral LD50 was determined to be > 2000 mg/kg bw.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11 May 2017 - 05 July 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Version / remarks:

2001 December 17th

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Version / remarks:

31 May 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI rats

Sex:

female

Details on test animals or test system and environmental conditions:

Source: TOXI COOP ZRT. Cserkesz u. 90., 1103 Budapest, Hungary
Hygienic level at arrival: SPF
Hygienic level during the study: Good conventional
Justification of strain: The Wistar rat as a rodent is one of the standard species of acute toxicity studies
Number of animals: 3 animals/group
Sex: Female, nulliparous and non pregnant animals
Age of animals: Young adult rat, 8 weeks old in the first and second step
Body weight range at starting (first step): 173 - 173 g
Body weight range at starting (second step): 197 - 200 g
Acclimatization time: 6 days in the first step and 7 days in the second step

Husbandry
Animal health: Only healthy animals were used for the study. Health status was certified by the study director.
Room: 13/2
Housing: Group caging (3 animals/cage)
Cage type: Type III. polypropylene/polycarbonate.
Bedding: Laboratory bedding.
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 22 ± 3 °C
Relative humidity: 30 - 70 %
Ventilation: above 10 air exchanges/hour by central air-condition system.
The temperature and relative humidity were recorded daily during the study.

Food and Water Supply
Animals received ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany and tap water from municipal supply, as for human consumption from bottle ad libitum. The diet and drinking water are periodically analysed and are considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

Route of administration:

oral: gavage

Vehicle:

vegetable oil

Remarks:

Helianthi annui oleum raffinatum

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

6 females/dose (3 females/step)

Control animals:

no

Details on study design:

Duration of observation period after the treatment: 14 days.

Frequency of observations:
Animals were observed individually after dosing at least once during the first 30 minutes, then 1 h, 2 h, 3 h, 4 h after the treatment and twice each
day for 14 days thereafter.

Body weight measurement:
The body weights were recorded on day 0 (just before the treatment), on day 7 and on day 15 with a precision of 1 g.

Necropsy:
At the end of the observation period all surviving rats were necropsied.

Statistics:

No statistics was used in the study.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No lethality was noted at a single oral dose of 2000 mg/kg bw.

Clinical signs:

other: In the first and second step, dark faeces were observed in all animals on Day 1, being related to the physical property of the test item. No systemic clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the

Gross pathology:

All organs of the animals treated with 2000 mg/kg bw proved to be free of treatment related gross pathological changes.

Summary of Lethality: Post-treatment observation period (14 days)

Groups	Treatment		Lethality
	Test Item	Dose (mg/kg bw)	Females
1	Step 1	2000	0/3
2	Step 2	2000	0/3

Summary of Clinical Symptoms

Groups	Treatment		Symptoms	Incidence
	Test Item	Dose (mg/kg bw)
1	Step 1	2000	Dark faeces	3/57
			Normal	54/57
2	Step 2	2000	Dark faeces	3/57
			Normal	54/57

 

Remark: Incidence = Number of symptoms/Summarized number of observations inside the group

Summarized number of observations inside the group = (number of observations of first animal) + (number of observations of second animal) +
(number of observations of third animal)

Summary of Body Weights (g)

FEMALES		Day 0	Day 7	Day 15
Group 1:
2000 mg/kg bw, Step 1
Group size:		3	3	3
Mean: (g)		173.0	206.7	243.3
SD:		0.00	4.73	20.50
FEMALES		Day 0	Day 7	Day 15
Group 2:
2000 mg/kg bw, Step 2
Group size:		3	3	3
Mean: (g)		198.3	226.3	238.7
SD:		1.53	8.08	12.50

Summary of Body Weight Gains (g)

 

 

FEMALES		Day 0-7	Day 7-15	Day 0-15
Group 1:
2000 mg/kg bw, Step 1
Group size:		3	3	3
Mean: (g)		33.7	36.7	70.3
SD:		4.73	15.95	20.50
FEMALES		Day 0-7	Day 7-15	Day 0-15
Group 2:
2000 mg/kg bw, Step 2
Group size:		3	3	3
Mean: (g)
SD:		28.0	12.3	40.3
		8.89	4.73	13.05

 

Interpretation of results:

GHS criteria not met

Conclusions:

The acute oral LD50 was determined to be > 2000 mg/kg bw.

Executive summary:

An acute oral toxicity study was performed according to OECD guideline 423. The acute toxic class method was carried out involving a stepwise procedure with the use of 2000 mg/kg bw as the starting dose in three female rats. No animal died in the first step at 2000 mg/kg bw dose level, so treatment with 2000 mg/kg bw was repeated on further three female rats. No animal died in the second step, too, so the test was finished, the stopping criteria of Annex 2d of OECD Guideline No. 423 were met. Animals were weighed, observed for lethality and toxic symptoms for 14 days after the treatment. Gross pathological examination was carried out on the 15th day after the treatment.

No lethality was noted at a single oral dose of 2000 mg/kg bw. In the first and second step, dark faeces were observed in all animals on Day 1, being related to the physical property of the test item. No systemic clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the general state and behaviour of the experimental animals were normal. The body weight development was not affected in all animals. All organs of the animals treated with 2000 mg/kg bw proved to be free of treatment related gross pathological changes. The method used is not intended to allow the calculation of a precise LD50 value. The acute oral LD50 was determined to be > 2000 mg/kg bw.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

2 000 mg/kg bw

Acute toxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Acute oral toxicity:

An acute oral toxicity study was performed according to OECD guideline 423. The acute toxic class method was carried out involving a stepwise procedure with the use of 2000 mg/kg bw as the starting dose in three female rats. No animal died in the first step at 2000 mg/kg bw dose level, so treatment with 2000 mg/kg bw was repeated on further three female rats. No animal died in the second step, too, so the test was finished, the stopping criteria of Annex 2d of OECD Guideline No. 423 were met. Animals were weighed, observed for lethality and toxic symptoms for 14 days after the treatment. Gross pathological examination was carried out on the 15th day after the treatment.

No lethality was noted at a single oral dose of 2000 mg/kg bw. In the first and second step, dark faeces were observed in all animals on Day 1, being related to the physical property of the test item. No systemic clinical symptoms were observed on the day of the treatment and during the 14-day observation period, the general state and behaviour of the experimental animals were normal. The body weight development was not affected in all animals. All organs of the animals treated with 2000 mg/kg bw proved to be free of treatment related gross pathological changes. The method used is not intended to allow the calculation of a precise LD50 value. The acute oral LD50 was determined to be > 2000 mg/kg bw.

Acute inhalation toxicity:

The study does not need to be conducted as exposure of humans via inhalation is not the appropriate route of exposure. Furthermore, physicochemical and toxicological properties do not suggest a potential for a significant rate of respiratory absorption.

Acute dermal toxicity:

According to REACH Annex VIII point 8.5.3 the study does not need to be conducted because the substance does not meet the criteria for classification as acute toxicity or STOT SE by the oral route and no systemic effects have been observed in in vivo studies with dermal exposure (e.g. skin irritation, skin sensitisation).

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data on acute oraltoxicity, the test item is not classified according to Regulation (EC) No 1272/2008 (CLP), as amended for the twelfth time in Regulation (EU) No 2019/521.