Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Between 20 September 2011 and 11 October 2011.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Deviations:
no
Principles of method if other than guideline:
The sequence of dosing may not always follow the Test Guideline as shown in the schematic diagram in attachment 1. It is Company Policy to minimisethe number of animals used on each study in accordance with UK Government Home Office guidelines.
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Details on test material:
Sponsor's identification: Benzenesulfonic acid, 4-C10-13-sec-alkyl derivs., compds. with N-[3-(dimethylamino)propyl]-2-methyl-2-propenamide (1:1)
Description : light amber coloured viscous liquid
Batch number: P1113
Purity : 96.3 w%
Date received: 01 July 2011
Expiry date : 30 March 2012
Storage conditions: room temperature in the dark

The integrity of supplied data relating to the identity, purity and stability of the test item is the responsibility of the Sponsor.

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
Female Wistar (RccHan:WIST) strain rats were used.
On receipt the animals were randomly allocated to cages. The animals were nulliparous and non-pregnant. After an acclimatisation period of at least five days the animals were selected at random and given a number unique within the study by indelible ink marking on the tail and a number written on a cage card. At the start of the study the animals were eight to twelve weeks of age. The bodyweights fell within an interval of ±20% of the mean initial bodyweight of the first treated group.

The animals were housed in groups of three in suspended solid floor polypropylene cages furnished with woodflakes. With the exception of an overnight fast immediately before dosing and for approximately three to four hours after dosing, free access to mains drinking water and food was allowed throughout the study.
The diet, drinking water and bedding were routinely analysed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.
The temperature and relative humidity were set to achieve limits of 19 to 25°C and 30 to 70% respectively. Any occasional deviations from these targets were considered not to have affected the purpose or integrity of the study. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (06:00 to 18:00) and twelve hours darkness.
The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on oral exposure:
All animals were dosed once only by gavage, using a metal cannula attached to a graduated syringe.
Doses:
2000 mg/kg
No. of animals per sex per dose:
3 female @ 2000 mg/kg
3 female @ 2000 mg/kg
Control animals:
no
Details on study design:
Procedure
Using available information on the toxicity of the test material, 2000 mg/kg was chosen as the starting dose.

Dose Level Concentration Dose Volume Number of Rats
(mg/kg) (mg/ml) (ml/kg) Female
2000 200 10 3
2000 200 10 3

All animals were dosed once only by gavage, using a metal cannula attached to a graduated syringe. The volume administered to each animal was calculated according to the fasted bodyweight at the time of dosing. Treatment of animals was sequential. Sufficient time was allowed between each group to confirm the survival of the previously dosed animals.
The animals were observed for deaths or overt signs of toxicity ½, 1, 2 and 4 hours after dosing and subsequently once daily for up to fourteen days.
Individual bodyweights were recorded prior to dosing and seven and fourteen days after treatment or at death.
At the end of the observation period the surviving animals were killed by cervical dislocation. All animals were subjected to gross pathological examination. This consisted of an external examination and opening of the abdominal and thoracic cavities for examination of major organs.
The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Results and discussion

Preliminary study:
Not applicable.
Effect levels
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: 95% confidence limits not reported
Mortality:
One animal was found dead one day after dosing.
Clinical signs:
Signs of systemic toxicity noted in one animal four hours after dosing were pilo-erection, hunched posture and noisy respiration. No signs of systemic toxicity were noted in surviving animals.
Body weight:
The surviving animals showed expected gains in bodyweight over the study period.
Gross pathology:
Abnormalities noted at necropsy of the animal that died during the study were abnormally red lungs, dark liver and dark kidneys. No abnormalities were noted at necropsy of animals that were killed at the end of the study.
Other findings:
None

Any other information on results incl. tables

Evaluation of Data

Data evaluations included the relationship, if any, between the exposure of the animal to the test item and the incidence and severity of all abnormalities including behavioural and clinical observations, gross lesions, bodyweight changes, mortality and any other toxicological effects.

Using the mortality data obtained, an estimate of the acute oral median lethal dose (LD50) of the test item was made as shown in the schematic diagram in Appendix 1 - (attachment 1).

Table 1              Individual Clinical Observations and Mortality Data

Dose Level mg/kg

Animal Number and Sex

Effects Noted After Dosing
(Hours)

Effects Noted During Period After Dosing
(Days)

½

1

2

4

1

2

3

4

5

6

7

8

9

10

11

12

13

14

2000

1-0

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

1-1

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

1-2

Female

0

0

0

PHRn

X

 

 

 

 

 

 

 

 

 

 

 

 

 

2-0

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

2-1

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

2-2

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0


0=     No signs of systemic toxicity

H =     Hunched posture

P =     Pilo-erection

Rn =   Noisy respiration

X =     Animal dead

Table 2              Individual Bodyweights and Weekly Bodyweight Changes

Dose Level

mg/kg

Animal Number and Sex

Bodyweight (g) at Day

Bodyweight (g)
at Death

Bodyweight Gain (g) During Week

0

7

14

1

2

2000

1-0 Female

162

178

188

 

16

10

1-1 Female

161

191

198

 

30

7

1-2 Female

165

-

-

147

-

-

2-0 Female

178

190

201

 

12

11

2-1 Female

188

195

216

 

7

21

2-2 Female

164

177

189

 

13

12

Table 3              Individual Necropsy Findings

Dose Level

mg/kg

Animal Number and Sex

Time of Death

Macroscopic Observations

2000

1-0 Female

Killed Day 14

No abnormalities detected

1-1 Female

Killed Day 14

No abnormalities detected

1-2 Female

Found dead Day 1

Lungs: abnormally red

Liver: dark

Kidneys: dark

2-0 Female

Killed Day 14

No abnormalities detected

2-1 Female

Killed Day 14

No abnormalities detected

2-2 Female

Killed Day 14

No abnormalities detected

Applicant's summary and conclusion

Interpretation of results:
Category 5 based on GHS criteria
Remarks:
Migrated information: >2000 - 5000 mg/kg
Conclusions:
The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2000 mg/kg bodyweight, with LD50 cut-off of 2500 mg/kg bodyweight (Globally Harmonised Classification System - Category 5 >2000 - 5000 mg/kg bodyweight).
Executive summary:

Introduction.

The study was performed to assess the acute oral toxicity of the test item following a single oral administration in the Wistar strain rat. The method was designed to be compatible with the following:

OECD Guidelines for the Testing of Chemicals No. 423 “Acute Oral Toxicity – Acute Toxic Class Method” (adopted 17 December 2001) Method B1tris Acute Toxicity (Oral) of Commission Regulation (EC) No. 440/2008

Method.

A group of three fasted females was treated with the test item at a dose level of 2000 mg/kg bodyweight. This was followed by a further group of three fasted females at the same dose level.

The test item was administered orally as asolutioninarachis oil BP. Clinical signs and bodyweight development were monitored during the study. All animals were subjected to gross necropsy.

Mortality. 

One animal was found dead one day after dosing.

Clinical Observations. 

Signs of systemic toxicity noted in one animal four hours after dosing were pilo-erection, hunched posture and noisy respiration. No signs of systemic toxicity were noted in surviving animals.

Bodyweight. 

The surviving animals showed expected gains in bodyweight over the study period.

Necropsy. 

Abnormalities noted at necropsy of the animal that died during the study were abnormally red lungs, dark liver and dark kidneys. No abnormalities were noted at necropsy of animals that were killed at the end of the study.

Conclusion. 

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2000 mg/kg bodyweight, with LD50 cut-off of 2500 mg/kg bodyweight (Globally Harmonised Classification System - Category 5 >2000 - 5000 mg/kg bodyweight).