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Acute Toxicity: oral

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Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
February 23 - March 30, 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report date:
2017

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
17th December 2001
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
acute toxic class method
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
N-propylphthalimide
EC Number:
226-189-1
EC Name:
N-propylphthalimide
Cas Number:
5323-50-2
Molecular formula:
C11H11NO2
IUPAC Name:
2-propyl-1H-isoindole-1,3(2H)-dione
Constituent 2
Chemical structure
Reference substance name:
N-sec-butylphthalimide
EC Number:
233-295-1
EC Name:
N-sec-butylphthalimide
Cas Number:
10108-61-9
Molecular formula:
C12H13NO2
IUPAC Name:
2-sec-butyl-1H-isoindole-1,3(2H)-dione
Constituent 3
Chemical structure
Reference substance name:
N-butylphthalimide
EC Number:
216-157-5
EC Name:
N-butylphthalimide
Cas Number:
1515-72-6
Molecular formula:
C12H13NO2
IUPAC Name:
2-butyl-1H-isoindole-1,3(2H)-dione
impurity 1
Reference substance name:
Unknown impurities.
Molecular formula:
Not available as unknown impurities.
IUPAC Name:
Unknown impurities.
1
Chemical structure
Reference substance name:
Water
EC Number:
231-791-2
EC Name:
Water
Cas Number:
7732-18-5
Molecular formula:
H2O
IUPAC Name:
water
Test material form:
liquid
Details on test material:
Batch CNAA053604
Color: clear yellowish
Form: liquid
Storage conditions: Room temperature in a closed container, protected from light.

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals or test system and environmental conditions:
Species and strain: Crl:WI rats
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 rats 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 first, second and third step
Body weight range
at starting (first step): 165 - 170 g
Body weight range
at starting (second step): 166 - 178 g
Body weight range
at starting (third step): 167 - 171 g
Acclimatization time: 5 days in first step, 6 days in second step and 7 days in third step

Housing: Group caging (3 animals/cage)
Cage type: Type II 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.

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.

In life phase: February 28- March 17, 2017


Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: Helianthi annui oleum raffinatum
Details on oral exposure:
All doses were formulated in the vehicle. Concentration of formulations was adjusted to maintain a treatment volume
of 10 mL/kg bw. The test item was applied in a concentration of 200 and 30 mg/mL. Formulations were prepared just
before the administration and were stirred continuously during the treatment.

Vehicle:
Name: Helianthi annui oleum raffinatum
Batch number: 1607-4569
Date of expiration: 30.11.2017
Produced by: Parma Produkt Kft.

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. All females died, so the test was continued at 300 mg/kg bw dose level on further three female rats.
No animal died in the second step at 300 mg/kg bw dose level, three further female rats were treated with the same (300 mg/kg bw) dose.
No animal died in the third step, too, so the test was finished, the stopping criteria of Annex 2d of OECD Guideline No. 423. was met.
Doses:
2000 mg/kg bw
300 mg/kg bw
No. of animals per sex per dose:
2000 mg/kg bw: 3 animals
300 mg/kg bw: 6 animals
Control animals:
no
Details on study design:
5 days in first step, 6 days in second step and 7 days in third step of acclimatization, treatment’s day,
14 days post-treatment observation period, necropsy on Day 15.

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 once each day for 14 days thereafter.

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

Necropsy:
All animals treated with 2000 mg/kg bw dose spontaneously died during the study and were necropsied on Day 1.
All animals treated with 300 mg/kg bw dose survived until the scheduled necropsy on Day 15.
Statistics:
No statistics was used in the study.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 300 - < 2 000 mg/kg bw
Based on:
test mat.
Mortality:
All rats dosed at 2000 mg/kg bw died on Day 1.
No death occurred at 300 mg/kg bw single oral dose of the test item.
Clinical signs:
2000 mg/kg bw: CNS - and emotion symptoms (decreased activity, bedding digging, closed eyes), disturbances of coordination
(incoordination, prone position), decreased righting reflex, decreased muscular tension (grip- and limb tone, body tone, abdominal tone)
and disturbances of the autonomic functions (dyspnea, redness skin and mucous membrane) were observed in animals between treatment day and Day 1.

2000 mg/kg bw: no clinical symptoms were observed on the day of the treatment and during the 14-day observation period.
Body weight:
The body weight development was undisturbed in all survivor animals.
Gross pathology:
All animals treated with 2000 mg/kg bw dose died spontaneously during the study and were necropsied on the Day 1.
All animals treated with 300 mg/kg bw dose survived until the scheduled autopsy on Day 15.
Autopsy revealed treatment related internal changes as haemorrhaged mucous membrane in the stomach, urinary bladder full of urine,
stomach full of gas and spotted liver in 2000 mg/kg bw dose group.
All organs of the animals treated with 300 mg/kg bw proved to be free of treatment related gross pathological changes.

Any other information on results incl. tables

A single oral administration - followed by a fourteen-day observation period - was performed by gavage.

The day before treatment the animals were fasted. The food but not water was withheld overnight.

Animals were weighed before the application and the food was given back 3 hours after the treatment.

Starting dose was selected on the basis of the available information about the test item.

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. All females died, so the test was continued at 300 mg/kg bw dose level

on further three female rats. No animal died in the second step at 300 mg/kg bw dose level, three further female rats

were treated with the same (300 mg/kg bw) dose.

No animal died in the third step, too, so the test was finished, the stopping criteria of Annex 2d

of OECD Guideline No. 423. was met.

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
The test item was ranked into classes of Globally Harmonized Classification System (GHS)
described in the OECD Guideline No. 423 as below:

Dose (mg/kg bw) 2000 300
Mortality (dead/treated) 3/3 0/6
LD50 (mg/kg bw) between 300 and 2000
GHS category 4

The estimated LD50 value is 500 mg/kg bw on basis of LD50 cut off mg/kg bw, because three animals died in 2000 mg/kg bw at 1st step.
Executive summary:

A single oral administration - followed by a fourteen-day observation period - was performed by gavage.

The day before treatment the animals were fasted. The food but not water was withheld overnight.

Animals were weighed before the application and the food was given back 3 hours after the treatment.

Starting dose was selected on the basis of the available information about the test item.

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. All females died, so the test was continued at 300 mg/kg bw dose level

on further three female rats. No animal died in the second step at 300 mg/kg bw dose level, three further female rats

were treated with the same (300 mg/kg bw) dose.

No animal died in the third step, too, so the test was finished, the stopping criteria of Annex 2d

of OECD Guideline No. 423. was met.

GHS category: 4

The estimated LD50value is 500 mg/kg bw on basis of LD50cut off mg/kg bw, because three animals died in 2000 mg/kg bw at 1ststep.