Bis{tris[4-(mono and dimethylamino)phenyl]methylium} 2-(bis{4-[ethyl(3-sulfonatobenzyl)amino]phenyl}methyliumyl)benzenesulfonate

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 2008/03/17 to 2008/04/02

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The reliability is rated 1 because the study followed the standard guideline of reference (OECD 423), which describes a procedure designed to evaluate this endpoint, the results were reviewed for reliability and assessed as valid, and the study was conducted under GLP condition.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Bis{tris[4-(mono and dimethylamino)phenyl]methylium} 2-(bis{4-[ethyl(3- sulfonatobenzyl)amino]phenyl}methyliumyl)benzenesulfonate

IUPAC Name:

Bis{tris[4-(mono and dimethylamino)phenyl]methylium} 2-(bis{4-[ethyl(3- sulfonatobenzyl)amino]phenyl}methyliumyl)benzenesulfonate

Constituent 2

Reference substance name:

Bis{tris[4-(mono and dimethylamino)phenyl]methylium} 2-(bis{4-[ethyl(3-sulfonatobenzyl)amino]phenyl}methyliumyl)benzenesulfonate

EC Number:

700-610-3

Molecular formula:

UVCB substance - not applicable

IUPAC Name:

Bis{tris[4-(mono and dimethylamino)phenyl]methylium} 2-(bis{4-[ethyl(3-sulfonatobenzyl)amino]phenyl}methyliumyl)benzenesulfonate

Details on test material:

- Name of test material : Sepisol Fast Blue 2BR
- Physical state/ appearance: dark blue powder
- Lot/batch No.: 905536
- Storage condition of test material: ambient temperature and in the dark

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: élevage JANVIER (53940 Le Genest-St-Isle, France).
- Age at study initiation: approximately 8 weeks.
- Weight at study initiation: At D1, mean (for group 1 and 2): 225.8 gram +/- 7.8.
- Fasting period before study: deprived of food overnight prior administration.
- Housing:Housed in cage (1 cm x 46 cm x 19 cm) with a stainless steel lid. Maximum 3 animals per cage. Sterilized and dust free sawdust litter.
- Diet : ad libitumexcept the night prior administration and 3-4 hours following administration (granules A04-10).
- Water: ad libitum. Distributed a propylene biberon with a stainless steel teat.
- Acclimation period:at least 5 days before dosing.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): 50 +/- 20
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 hrs/12hrs

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

VEHICLE
- Concentration in vehicle: adapted for each animal
- Amount of vehicle : 5 mL/kg
- Justification for choice of vehicle: corn oil was chosen among various vehicules because it does not induce pain. Following several preliminary assays, the aqueous-based vehicles (pure water, HCMC) were disregarded and the non-polar vehicle (corn oil commonly used for oral toxicity assay ) has been chosen since it allowed to prepare a homogenous mixture usable for oral adiministration.
- Lot/batch no. : 07030169/C

Doses:

2000 mg/kg for both groups.

No. of animals per sex per dose:

3 rats for both groups (6 in total)

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations: regularly following administration (immediately, 30 min, 1h, 2h, 3h and 4 h) and at least once a day until the end of the study
- Frequency of weighing: D-1, D1 T0, D4, D8, D15
- Necropsy of survivors performed: yes

Results and discussion

Preliminary study:

first dose of 2000 mg/kg with 3 animals.

Effect levels

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

At the 2000 mg/Kg dose: no deaths were observed.

Clinical signs:

other: At the 2000 mg/kg dose, significant symptoms occured: important piloerection and reduced motor activity which persisted 2-3 days after administration of the test item. Colored feces on D1. No organic lesion or tissue injury observed at the autopsy.

Any other information on results incl. tables

Dose = 2000 mg/kg

Clinical observations

Time of observation	Comments	Time of observation	Comments
D1 (following administration)	Important piloerection for all animals and reduced motor activity for 2 of them.	D6	Nothing to report
D1 T30'	Important piloerection and reduced motor activity  for all animals.	D7	Nothing to report
D1 T1h	Similar to D1 T30' and blue feces.	D8	Nothing to report
D1 T2h	Similar to D1 T30'.	D9	Nothing to report
D1 T3h	Similar to D1 T30'.	D10	Nothing to report
D1 T4h	Similar to D1 T30'.	D11	Nothing to report
D2	Similar to D1 T30'.	D12	Nothing to report
D3	For 3 animals:  similar to D1 T30'. Slight piloerection and reduced motor activity for the other 3.	D13	Nothing to report
D4	Important piloerection and round back for 1 animal. Nothing to report for all other animals.	D14	Nothing to report
D5	Nothing to report	D15	Nothing to report

Autopsy

Animals N°		Mortalilty		Comments
		Day	Cause
1^stStep	5632	D15	Euthanasia	Death by sacrifice
	5633	D15	Euthanasia	Death by sacrifice
	5634	D15	Euthanasia	Death by sacrifice
2^dStep	5635	D15	Euthanasia	Death by sacrifice
	5636	D15	Euthanasia	Death by sacrifice
	5637	D15	Euthanasia	Death by sacrifice

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

According to the CLP regulation, the test material Sepisol Fast Blue 2BR has not been classified as acute toxicity with a
LD 50 determined to be above the threshold of 2000 mg/kg bw.

Executive summary:

The aim of this study was to assess qualitatively and quantitatively the toxic effects and the delay of the appearance after single oral administration of a pre-defined dose of 2000 mg/kg body weight, of test element named SEPISOL FAST BLUE 2BR suspended in corn oil, in 6 females rats, using a stepwise procedure. The study has been performed by using the OECD guideline 423.

The animals were daily observed for at least 14 days after administration and the clinical signs and signs of toxicity were noted.

A preliminary test was performed with a 2000 mg/kg dose after which no animals dies.

The results of the assay showed that 6 animals did not die with the 2000 mg/kg dose level.

Significant symptoms occured: important piloerection, reduced motor activity which persisted 2-3 days after administration.

No organic lesion or tissue injury has been observed at the autopsy.

This test provided results allowing the test element not to be classified according to the Classification and Labeling of Products regulation (CLP EC 1272/2008).