Reactive Orange F08-0314

Acute Toxicity: dermal

Administrative data

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 December 2010 to 29 December 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted to EU, OECD and US EPA 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 guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1200 (Acute Dermal Toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

yes

Test material

Test material information

Constituent 1

Details on test material:

Product name: Reactive Orange F08-0314

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Species and strain: RjHan:(WI) Wistar rats
Source: Laboratoire Elevage Janvier, B.P. 4105, Route des Chênes Secs, 53940 Le Genest-St-Isle CEDEX FRANCE
Hygienic level at arrival: SPF
Hygienic level during the study: Standard housing conditions
Justification of strain: The Wistar rat is one of the standard rodent species used in acute toxicity studies
Number of animals: 5 animals/sex
Sex: Male and female, female rats were nulliparous and non-pregnant.
Age of animals at study start: Young adult rats
Body weight range at dosing: Between 210 g and 249 g
Acclimatization time: 5 days
Animal health: Only healthy animals were used for the study. The veterinarian certified the health status.
Room-Box: 242/6
Housing: Individual caging
Cage type: Type II. polypropylene/polycarbonate
Bedding: Laboratory bedding: Lignocel Hygienic Animal Bedding produced by J. Rettenmaier & Söhne GmbH+Co.KG (Holzmühle 1, 73494 Rosenberger, Germany);

Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 20.4- 25.0 °C
Relative humidity: 24 - 64 %
Ventilation: 15-20 air exchanges/hour
Enrichment: Rodents were housed with deep wood sawdust bedding to allow digging and other normal rodent activities.

The temperature and relative humidity was 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 not considered 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). The quality control results are retained in the archives at LAB Research Ltd.

Identification
The individual identification was performed using numbers written on the tail with a 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 containing information about study code, sex, dose group, cage number and individual animal numbers.

Administration / exposure

Type of coverage:

semiocclusive

Vehicle:

water

Details on dermal exposure:

The back of each animal was shaved (approximately 10 % area of the total body surface) approximately 24 hours prior to treatment. The test item was applied as a single dose as supplied to the shaved skin and remained in contact with the skin for the 24- hour exposure period. For that purpose, the appropriate amount of the test item was moistened with distilled water and distributed as uniformly as possible. Sterile gauze pads were placed on the skin of rats to cover the test item. These gauze pads were kept in contact with the skin by a patch with adhesive hypoallergenic plaster. The entire trunk of the animal was then wrapped with semi occlusive plastic wrap for 24 hours.

At the end of the exposure period, the area of skin treated with the test item was washed with water of body temperature.

Duration of exposure:

24 hours

Doses:

The test item was not expected to be lethal at 2000 mg/kg bw. A limit test was therefore performed.

No. of animals per sex per dose:

5 males/5 females per dose

Control animals:

no

Details on study design:

Clinical Observations
Clinical observations were performed on the day of treatment at 1 and 5 hours after application of the test item and once each day for 14 days thereafter. Observations included the skin and fur, eyes and mucous membranes, the 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.

Measurement of Body Weight
The body weights were recorded on Day 0 (before test item administration) and on Days 3, 7 and 14.

NECROPSY
All animals were anaesthetised with Euthasol®40% and exsanguinated. After examination of the external appearance, the cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs was observed. All macroscopic changes were recorded.

Statistics:

No data

Results and discussion

Preliminary study:

Not applicable

Effect levels

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred after a 24-hour dermal exposure of Reactive Orange F08-0314 administered at 2000 mg/kg bw to RjHan:(WI) Wistar rats followed by a 14-day observation period.

Clinical signs:

other: No clinical signs were observed after the treatment with the test item or during the 14 day observation period.

Gross pathology:

A single 24 hour dermal application of Reactive Orange F08-0314 to the Rj Han: (WI) rat at a dose level of 2000 mg/kg bw was not accompanied with any macroscopic test item-related findings.

Other findings:

Orange staining was recorded on the skin in all animals after dosing. The discoloration of the skin lasted up to 13 days in all animals. No other local dermal signs were observed.

Any other information on results incl. tables

CLINICAL OBSERVATIONS

INDIVIDUAL CLINICAL OBSERVATIONS

Study Code: 10296-002P              Test System: RjHan: WI Rat        Test Item Reactive Orange F08-0314

DOSE LEVEL: 2000 mg/kg

Cage No.	Animal No./Sex	Observations	Observation days																Frequency
			0		1	2	3	4	5	6	7	8	9	10	11	12	13	14
			1h	5h
1	3403 Male	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	+	+	+	+	6/16
			-	-	+	+	+	+	+	+	+	+	+	+	-	-	-	-	10/16
2	3404 Male	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	+	+	+	+	6/16
			-	-	+	+	+	+	+	+	+	+	+	+	-	-	-	-	10/16
3	3405 Male	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	+	+	+	+	6/16
			-	-	+	+	+	+	+	+	+	+	+	+	-	-	-	-	10/16
4	3406 Male	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	+	+	+	+	6/16
			-	-	+	+	+	+	+	+	+	+	+	+	-	-	-	-	10/16
5	3407 Male	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	+	+	+	+	6/16
			-	-	+	+	+	+	+	+	+	+	+	+	-	-	-	-	10/16
6	3408 Female	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	-	-	+	+	4/16
			-	-	+	+	+	+	+	+	+	+	+	+	+	+	-	-	12/16
7	3409 Female	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	-	-	+	+	4/16
			-	-	+	+	+	+	+	+	+	+	+	+	+	+	-	-	12/16
8	3410 Female	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	-	-	-	+	3/16
			-	-	+	+	+	+	+	+	+	+	+	+	+	+	+	-	13/16
9	3411 Female	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	-	-	+	+	4/16
			-	-	+	+	+	+	+	+	+	+	+	+	+	+	-	-	12/16
10	3512 Female	Symptom Free Orange skin	+	+	-	-	-	-	-	-	-	-	-	-	+	+	+	+	6/16
			-	-	+	+	+	+	+	+	+	+	+	+	-	-	-	-	10/16

Remarks:            +: Present

                               h=hour(s)           Treatment day = Day 0

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

 

BODY WEIGHT DATA

INDIVIDUAL BODY WEIGHT AND BODY WEIGHT GAIN

Study Code: 10/296-002P            Test System: Rj Han: WI Rat       Test Item: Reactive Orange F08-0314

DOSE LEVEL: 2000 mg/kg

Cage No.	Animal No. / Sex	Body Weight (g) Days				Body Weight Gain (g)
		0	3	7	14	0-3	3-7	7-14	0-14
1	3403 Male	249	260	295	348	11	35	53	99
2	3404 Male	240	258	289	334	18	31	45	94
3	3405 Male	232	254	278	328	22	24	50	96
4	3406 Male	243	260	282	330	17	22	48	87
5	3407 Male	242	266	292	344	24	26	52	102
Mean:		241.2	259.6	287.2	336.8	18.4	27.6	49.6	95.6
Standard deviation:		6.1	4.3	7.0	8.8	5.0	5.3	3.2	5.7
6	3408 Female	215	219	225	237	4	6	12	22
7	3409 Female	222	228	245	261	6	17	16	39
8	3410 Female	210	217	227	243	7	10	16	33
9	3411 Female	223	226	237	250	3	11	13	27
10	3412 Female	226	231	237	253	5	6	16	27
Mean:		219.2	224.2	234.2	248.8	5.0	10.0	14.6	29.6
Standard deviation:		6.5	6.0	8.2	9.2	1.6	4.5	1.9	6.5

Remarks:            Treatment day – Day 0

 

MACROSCOPIC FINDINGS

INDIVIDUAL INTERNAL AND EXTERNAL MACROSCOPIC OBSERVATIONS

Study Code: 10/296-002P            Test System: Rj Han: WI Rat       Test Item: Reactive Orange F08-0314

DOSE LEVEL: 2000 mg/kg

Cage No.	Animal No. / Sex	Necropsy Date	External Observations	Internal Observations	Organ/Tissue
1	3403 Male	29 December 2010	No external observations	No internal observations	Not applicable
2	3404 Male	29 December 2010	No external observations	No internal observations	Not applicable
3	3405 Male	29 December 2010	No external observations	No internal observations	Not applicable
4	3406 Male	29 December 2010	No external observations	No internal observations	Not applicable
5	3407 Male	29 December 2010	No external observations	No internal observations	Not applicable
6	3408 Female	29 December 2010	No external observations	No internal observations	Not applicable
7	3408 Female	29 December 2010	No external observations	No internal observations	Not applicable
8	3410 Female	29 December 2010	No external observations	No internal observations	Not applicable
9	3411 Female	29 December 2010	No external observations	No internal observations	Not applicable
10	3412 Female	29 December 2010	No external observations	In estrus	Uterus

 

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The acute dermal median lethal dose (LD50) of the test item Reactive Orange F08-0314 was found to be higher than 2000 mg/kg body weight in male and female RjHan:(WI) Wistar rats.

Executive summary:

An acute dermal toxicity study was performed with test item Reactive Orange F08 -0314 in RjHam;(WI) Wistar rats, in compliance with OECD Guideline No.: 402 (24th Feb. 1987), Commission Regulation (EC) No 440/2008, B.3 (L 142, 30 May 2008) and OPPTS 870.1200 (EPA 712 -C-98 -192, August 1998). This study is compliant with the Principles of Good Laboratory Practice (GLP) and reported with a valid GLP certificate.

A limit test was carried out at 2000 mg/kg body weight (bw) in both sexes (5 rats/sex). The test item was applied as supplied, moistened with distilled water, as a single dermal 24-hour exposure followed by a 14‑day observation period.

Clinical observations were performed on all animals at 1 and 5 hours after dosing and daily for 14 days thereafter. Body weight was measured prior to dosing on Day 0 and on Days 3, 7 and 14.Ratswere euthanized and a gross macroscopic examination performed at the end of the 2-week observation period (Day 14).

 

The results of the study were summarized as follows:

 

Mortality

No mortality occurred.

 

Systemic clinical signs

No clinical signs were observed after the treatment with the test item or during the 14‑day observation period.

 

Local dermal signs

No local dermal signs were observed during the entire study period. However, orange staining was observed on the skin in all animals after dosing from Day 1 to Day 13.

 

Body weight

The body weight and body weight gain of Reactive Orange F08-0314 treated animals did not show any test item-related effect.

 

Necropsy

There was no evidence of test item-related observations at a dose level of 2000 mg/kg at necropsy. The uterus in estrus was seen in the female 3412.

 

Conclusions

The acute dermal median lethal dose (LD₅₀) of the test itemReactive Orange F08-0314was found to be higher than 2000 mg/kg bw in male and female RjHan:(WI) Wistar rats.