Hydrogen 9-amino-7-phenyl-5-(phenylamino)-4,10-disulphonatobenzo[a]phenazinium, disodium salt

Acute Toxicity: dermal

Administrative data

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20 December 2017 to 02 February 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

OECD Guideline for Testing of Chemicals No. 402 (Section 4: Health Effects) “Acute Dermal Toxicity: Fixed Dose Procedure” adopted on 09 October 2017

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Hydrogen 9-amino-7-phenyl-5-(phenylamino)-4,10-disulphonatobenzo[a]phenazinium, disodium salt

EC Number:

276-817-3

EC Name:

Hydrogen 9-amino-7-phenyl-5-(phenylamino)-4,10-disulphonatobenzo[a]phenazinium, disodium salt

Cas Number:

72749-80-5

Molecular formula:

C28H20N4O6S2.2Na

IUPAC Name:

Benzo[a]phenazinium, 9-amino-7-phenyl-5-(phenylamino)-4,10-disulfo-, inner salt, sodium salt (1:2)

Test material form:

solid: particulate/powder

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: In-house bred animals
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 9 to 10 weeks
- Weight at study initiation: 220.21 g to 232.16 g
- Fasting period before study: No
- Housing: L 430 x B 285 x H 150 mm
- Diet (e.g. ad libitum): Altromin maintenance diet for rats and mice (manufactured by Altromin Spezialfutter GmbH & Co. KG)
- Water (e.g. ad libitum): Deep bore-well water passed through Reverse osmosis unit
- Acclimation period: Start: 20 December 2017 End: 01 January 2018

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.7°C to 23.0°C
- Humidity (%): 48% to 67%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light and 12 hours dark cycle

IN-LIFE DATES: From: 20 December 2017 To: 11 January 2018

Administration / exposure

Type of coverage:

semiocclusive

Vehicle:

water

Details on dermal exposure:

TEST SITE
- Area of exposure: approximately 10% of the total body surface
- % coverage: approximately 10% of the total body surface
- Type of wrap if used: non-irritating adhesive tape

REMOVAL OF TEST SUBSTANCE
- Washing (if done): distilled water
- Time after start of exposure: 24 hours

TEST MATERIAL
- For solids, paste formed: yes

VEHICLE
- Amount(s) applied (volume): 0.5 mL

Duration of exposure:

The contact period of test item was at least 24 hours

Doses:

2000 mg/kg body weight

No. of animals per sex per dose:

range finding study: 1 female (2000 mg/kg body weight)
main study: 2 females (2000 mg/kg body weight)

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: once daily for clinical signs of toxicity and twice daily for mortality and body weights weekly once
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight

Results and discussion

Effect levels

Key result

Sex:

female

Dose descriptor:

LD0

Effect level:

>= 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortalities observed

Clinical signs:

other: No treatment related clinical signs were observed

Gross pathology:

No treatment related gross pathological changes were observed

Any other information on results incl. tables

Phase of the Experiment

Dose (mg/kg body weight)

Animal No.

Sex

Time of Dosing (AM)

Clinical Signs of Toxicity and Mortality on Day 1

Clinical Signs of Toxicity and Mortality on days

20-30 mins

1 hr (±10 mins)

2 hrs (±10 mins)

4 hrs (±10 mins)

6 hrs (±10 mins)

2

3

4

5

6

7

8

9

10

11

12

13

14

15

Range Finding Study

2000

Rc6201

F

11:14

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Main Study

2000

Rc6202

F

11:16

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rc6203

F

11:18

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

    N: Normal; F: Female; min: minutes; hr/hrs: hour/hours

Phase of the Experiment	Dose (mg/kg body weight)	Animal No.	Sex	Body Weight (g) on Days				Percent Change in Body Weight with Respect to Day
				1	8	15		1 to 8	1 to 15
Range Finding Study	2000	Rc6201	F	242.41	265.76	297.45		9.63	22.71
Main Study	2000	Rc6202	F	232.78	261.75	290.45		12.45	24.77
		Rc6203	F	246.70	272.59	301.63		10.49	22.27
		Mean		239.74	267.17	296.04		11.47	23.52
		±SD		9.84	7.67	7.91		1.38	1.77

     F: Female; SD: Standard Deviation

Phase of the Experiment	Dose (mg/kg body weight)	Animal No.	Sex	Fate	Gross Pathology Findings
					External	Internal
Range finding Study	2000	Rc6201	F	TS	NAD	NAD
Main Study	2000	Rc6202	F	TS	NAD	NAD
		Rc6203	F	TS	NAD	NAD

 NAD: No Abnormality Detected; F: Female;TS: Terminal Sacrifice             

Applicant's summary and conclusion

Interpretation of results:

Category 5 based on GHS criteria

Conclusions:

Under the experimental conditions employed and based on the above results, it is concluded that the acute dermal median lethal dose (LD50) of test item Acid Violet 50 in Sprague Dawley rats is >2000 mg/kg body weight

Executive summary:

The test item Acid Violet 50 was evaluated for Acute Dermal Toxicity in Sprague Dawley Rats as per the OECD guideline for the testing of chemicals No. 402, “Acute Dermal Toxicity - Fixed Dose Procedure”.

The study was performed in two phases i.e. range finding study and main study. Range finding study was performed with one female rat and main study was performed with two female rats. On the day before the application of the test item, fur on the dorso-lateral area of the trunk of the animals was removed by clipping closely with an electric hair clipper and care was taken to avoid abrading the skin.

The required quantity of the test item was moistened with 0.5 mL of distilled water to form thin paste using glass rod and applied as uniform film over an area of approximately 10% of the total body surface. The test item was held on to the applied surface by covering with cotton gauze dressing and wrapped with non-irritating adhesive tape and finally the application site was wrapped using semi-occlusive crepe bandage. The contact period of test item was 24 hours. At the end of the contact period, the residual test item was washed using distilled water and dried with absorbent cotton.

The animals were dosed in a stepwise procedure with one female at a time in range finding study. Based on sponsor’s specification, a starting dose of 2000 mg/kg body weight was selected from the fixed dose levels of 50, 200, 1000 and 2000 mg/kg body weight. No clinical signs and mortalities were observed at the dose level of 2000 mg/kg body weight in range finding study. Hence, during main study, two animals were administered with the same dose level of 2000 mg/kg body weight. No clinical signs and mortalities were observed at the dose level of 2000 mg/kg body weight. Hence, no further testing was carried out.

All the animals were observed for clinical signs of toxicity and mortality at 20 to 30 min, 1 hr (±10 mins), 2 hrs (±10 mins), 4 hrs (±10 mins) and 6 hrs (±10 mins) post dosing on day 1 and thereafter once daily for clinical signs of toxicity and twice daily for mortality during the 14 days observation period. The body weight was recorded on day 1 before test item application and on day 8 and 15. At the end of observation period, all the animals were sacrificed under carbon dioxide anaesthesia and subjected to necropsy and detailed gross pathological examination.

No mortality, clinical signs and skin reactions were noted. No treatment related changes in body weight and percent change in body weight with respect to day 1 were noted. Normal increase in body weights were noted during the observation period. No treatment related gross pathological changes were noted at 2000 mg/kg body weight (range finding study and main study) during necropsy