Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
26 October 2010 to 10 November 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted to EU & OECD test guidance in compliance with GLP and reported with a valid GLP certificate.

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:
EU Method B.1 (Acute Toxicity (Oral))
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Name: Reactive Red F08-0146

Test animals

Species:
rat
Strain:
other: Wistar RjHan:WI
Sex:
female
Details on test animals and environmental conditions:
Species and strain: RjHan:WI rats
Source: Laboratoire Elevage Janvier, B.P. 4105, Route des Chênes Secs, 53940 Le Genest-St-Isle CEDEX FRANCE
Hygienic level at supplier: SPF
Hygienic level during the study: Standard housing conditions
Number of animals: 6 animals, 3 animals/group
Sex: Female, nulliparous and non-pregnant.
Age of animals at dosing: Young healthy adult rats, 8-10 weeks old
Date of receipt: 21 October 2010
Body weight at treatment: 197 – 214 g
Acclimation period: At least 5 days
Animal health: Only healthy animals were used for the test. The veterinarian certified health status.
Number of animal room: 522/2
Housing: 3 animals / cage
Cage type: Type III polypropylene/polycarbonate
Bedding: Lignocel Bedding for Laboratory Animals was available to animals during the study.
Lighting period: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 22 ± 3°C
Relative humidity: 30 - 70 %
Ventilation: 15-20 air exchanges/hour
Enrichment: Animals were housed by group to allow social interaction and with deep wood sawdust bedding to allow digging and other normal rodent activities.

The temperature and relative humidity were recorded twice daily during the study.

Food and Water Supply: Animals received ssniff® SM R/M-Z+H "Autoclavable complete feed for rats and mice – breeding and maintenance" produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany ad libitum, and tap water from the municipal supply, as for human consumption from 500 ml bottle ad libitum. The food is considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

Water quality control analysis is performed once every three months and microbiological assessment is performed monthly, by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József A.u.36., Hungary).

Animal Identification: Animals were individually identified using numbers written on the tail with an indelible marker pen. The numbers were given on the basis of LAB Research Ltd.' s Master File, for each animal allocated to the treatment groups. The cages were identified by cards, with information about study code, sex, dose group, cage number and individual animal numbers.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
Vehicle: Distilled water
Batch number: 3590210
Expiry Date: February 2013
Produced by: Teva zRt.
Dose volume: 10 mL/kg bw
Test item was freshly formulated at a concentration of 200 mg/mL in the vehicle on the day of administration. The formulation container was stirred continuously during administration with the objective of ensuring that the syringe was filled from a homogenous liquid.
Doses:
Initially, three female animals were treated with 2000 mg/kg bw of Reactive Red F08-0146. No mortality was observed, therefore further 3 animals were treated at the dose level of 2000 mg/kg bw. As no mortality was observed in this second dose group, further testing was not required according to the test guidelines.
No. of animals per sex per dose:
3 initial; 3 additional.
Control animals:
no
Details on study design:
The initial dose level was selected by the study director to be that which is most likely to produce mortality in some of the dosed animals. 2000 mg/kg bw was selected to be the starting dose.

A single oral gavage administration was followed by a fourteen-day observation period. On the day before treatment, the animals were fasted. The food but not water was withheld during an overnight period. Animals were weighed just before treatment. The test item was administered by oral gavage in the morning. The food was returned 3 hours after the treatment.

Clinical Observations
Clinical observations were performed on all animals at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter. Individual observations were performed on the skin, fur, eyes, mucous membranes, respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

Body Weight Measurement
The body weight was recorded on the day before treatment (Day -1), on the day of the treatment (day 0) and weekly after. Additional body weight measurement was carried out on Day 3.

Necropsy
Macroscopic examination was performed on all animals. The animals were sacrificed by exsanguination under pentobarbital anaesthesia (Euthasol® 40 %). After examination of the external appearance, the cranial, thoracic and the abdominal cavities were opened and the organs and the tissues were observed. Macroscopic abnormalities were recorded.
Statistics:
No data

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.
Mortality:
Reactive Red F08-0146 did not cause mortality at a dose level of 2000 mg/kg bw.
Clinical signs:
Treatment with Reactive Red F08-0146 caused coloured urine on Day 1 and coloured faeces on Day 0.
Body weight:
Body weight gains of Reactive Red F08-0146 treated animals during the study showed no indication of a treatment-related effect.
Gross pathology:
A single oral gavage of Reactive Red F08-146 to the RjHan:WI rat at a dose level of 2000 mg/kg bw, followed by a 14 day observation period, was not associated with any test item-related gross findings.
Other findings:
Urine & feaces coloured red in litter Day 0 & 1.

Any other information on results incl. tables

CLINICAL OBSERVATIONS

DOSE LEVEL: 2000 mg/kg bw                                                                                       SEX: FEMALE

Cage No.

Animal Number

Observations

Observation days

Frequency

0

1-14

30’

1h

2h

3h

4h

6h

1*

2119

Symptom Free

+

+

+

+

+

+

+

20/20

2120

Symptom Free

+

+

+

+

+

+

+

20/20

2121

Symptom Free

+

+

+

+

+

+

+

20/20

2**

2122

Symptom Free

+

+

+

+

+

+

+

20/20

2123

Symptom Free

+

+

+

+

+

+

+

20/20

2124

Symptom Free

+

+

+

+

+

+

+

20/20

Remarks:            +: present

                           h=hour (s)          Treatment day = Day 0

                           Frequency of observations = number of occurrence of observation / total number of observations

                            *: Urine coloured in litter (red) Day 1

                            Faeces coloured in litter (red) Day 0

                            **: Urine coloured in litter (red) Day 1

                            Faeces coloured in litter (red) Day 0

 

BODY WEIGHT DATA

DOSE LEVEL: 2000 mg/kg bw                                                                                      SEX: FEMALE

Cage No.

Animal No.

Body weight (g) Days

Body weight gain (g)

-1

0

3

7

14

1-10

0-3

3-7

7-14

-1-14

1

2119

216

212

233

250

260

-4

21

17

10

44

2120

219

210

227

234

257

-9

17

7

23

38

2121

222

214

234

252

269

-8

20

18

17

47

2

2122

217

202

224

242

260

-15

22

18

18

43

2123

209

197

219

225

237

-12

22

6

12

28

2124

213

204

220

223

239

-9

16

3

16

26

Mean:

216.0

206.5

226.2

237.7

253.7

-9.5

19.7

11.5

16.0

37.7

Standard deviation:

4.6

6.6

6.4

12.4

12.8

3.7

2.6

6.9

4.6

8.8

Remark:              Treatment day = Day 0

NECROPSY FINDINGS

DOSE LEVEL: 2000 mg/kg bw                                                                                                SEX: FEMALE

Cage No.

Animal ID

External Observations

Internal Observations

Organ/Tissue

1

2119

No external observations recorded

In estrus

Uterus

2120

No external observations recorded

In estrus

Uterus

Brown focus, few, all lobes

Lungs

2121

No external observations recorded

Brown focus, many, all lobes

Lungs

2

2122

No external observations recorded

In estrus

Uterus

2123

No external observations recorded

No internal observations recorded

Not applicable

2124

No external observations recorded

In estrus

Uterus

 

Applicant's summary and conclusion

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the conditions of this study, the acute oral LD50 value of the test item Reactive Red F08-0146 was found to be above 2000 mg/kg bw in female RjHan: WI rats.

According the GHS criteria, Reactive Red F08-0146 should be ranked as "Category 5" or "Unclassified".
Executive summary:

The single-dose oral toxicity of Reactive Red F08-0146 was performed according to the acute toxic class method (OECD 423 and Commission Regulation (EC) No 440/2008,B.1.tris) in RjHan: WI rats in compliance with the Principles of Good Laboratory Practice (GLP) and reported with a valid GLP certificate.

 

Two groups of three female RjHan:WI rats were treated with Reactive Red F08-0146 at a dose level of 2000 mg/kg bw (Group 1 and Group 2).

 

Initially, three females (Group 1) were treated at a dose level of 2000 mg/kg bw. As no mortality was observed, a confirmatory group (Group 2) was treated at the same dose level. No mortality was observed in the confirmatory group, therefore no further testing was required according to OECD 423 and Commission Regulation (EC) No 440/2008,B.1.tris.

 

A single oral treatment was carried out by gavage for each animal after an overnight food withdrawal. Food was made available again 3 hours after the treatment. Reactive Red F08-0146 was administered as a solution prepared in distilled water at a concentration of 200 mg/ml at a dosing volume of 10 ml/kg bw.

 

Clinical observations were performed at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter. Body weight was measured on Days -1, 0, 3, 7 and before necropsy. All animals were subjected to a necropsy and a macroscopic examination.

 

Results

Mortality: Reactive Red F08-0146 did not cause mortality at a dose level of 2000 mg/kg bw.

Clinical observations: Treatment with Reactive Red F08-0146 caused coloured urine on Day 1 and coloured faeces on Day 0.

Body weight and body weight gain: Body weight gains o fReactive Red F08-0146 treated animals during the study showed no indication of a treatment-related effect.

Macroscopic Findings: A single oral gavage of Reactive Red F08-146 to the RjHan:WI rat at a dose level of 2000 mg/kg bw, followed by a 14 day observation period, was not associated with any test item-related gross findings.

 

Conclusion

Under the conditions of this study, the acute oral LD50 value of the test item Reactive Red F08-014 6was found to be above 2000 mg/kg bw in female RjHan: WI rats.

 

According the GHS criteria, Reactive Red F08-0146 should be ranked as "Category 5" or "Unclassified".