Reaction mass of 5,5'-{(phenylmethanediyl)bis[benzene-4,1-diyl-diazene-2,1-diyl]}bis{1-[3-(dimethylamino)propyl]-4-methyl-6-oxo-3-(pyridinium-1-yl)-1,6-dihydropyridin-2-olate} hydrochloride and 5,5’-[3,4’-(phenylmethanediyl)diphenylene]bis(diazene-2,1-diyl)bis{1-[3-(dimethylamino)propyl]-4-methyl-6-oxo-3-(pyridinium-1-yl)-1,6-dihydropyridin-2-olate} hydrochloride

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

2007-09-24 to 2008-01-28

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline-conform study under GLP without deviations, conducted with the analogue substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

analoge substance (refer to IUCLID chapter 13)

IUPAC Name:

analoge substance (refer to IUCLID chapter 13)

Test animals

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd, Laboratory Animal Services, Wölferstrasse 4, 4414 Füllinsdorf / Switzerland
- Age at study initiation: 11 weeks
- Weight at study initiation: 184.3 - 203.5 g
- Fasting period before study: treatment applied after being fasted for approximately 17 to 18 hours (access to water was permitted). Food was provided again approximately 3 hours after dosing.
- Housing: In groups of three in Makrolon type-4 cages with wire mesh tops and standard softwood bedding
- Diet (e.g. ad libitum): Pelleted standard rat/mouse maintenance diet ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 30-70 %
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours dark

IN-LIFE DATES: From: 2007-09-21 To: 2007-10-11

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 200 and 30 mg/l, resp.
- Amount of vehicle (if gavage): 10 ml/kg
- Justification for choice of vehicle: Purified water was found to be a suitable vehicle. The vehicle was chosen after a non-GLP solubility trial which was performed before the study initiation date.
- Lot/batch no. (if required):
- Purity: Purified water prepared at RCC Ltd (deionised water which was processed and treated by the PURELAB Option-R unit. This latter links four purification technologies: reverse osmosis, adsorption, ion-exchange and photo oxidation).

MAXIMUM DOSE VOLUME APPLIED: 2.035 ml

DOSAGE PREPARATION (if unusual):
The dose formulations were made shortly before each dosing occasion using a magnetic stirrer, as homogenizer. The test item was first ground with a pestle and a mortar. The pulverized test item was then weighed into a tared glass beaker on a suitable precision balance and the vehicle was added (weight:volume). Homogeneity of the test item in the vehicle was maintained during administration using a magnetic stirrer.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: no data

Doses:

300 and 2000 mg/kg bw

No. of animals per sex per dose:

6 females in dose group 300 mg/kg bw
3 females in dose group 2000 mg/kg bw

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
Mortality / Viability: Daily during the acclimatization period, during the first 30 minutes and at approximately 1, 2, 3 and 5 hours after administration on test day 1 (with the clinical signs) and twice daily during days 2-15.
Body Weights: On test days 1 (prior to administration), 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed:
Clinical signs: Daily during the acclimatization period, during the first 30 minutes and at approximately 1, 2, 3 and 5 hours after administration on test day 1. Once daily during days 2-15. All abnormalities were recorded.
Further: body weight,organ weights

Statistics:

No statistical analysis was used.

Results and discussion

Effect levels

Sex:

female

Dose descriptor:

LD50

Effect level:

> 300 - < 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: Analogue substance.

Mortality:

MORTALITY:
The following animals were treated and percentage of mortality was observed:
3 females treated at 2000 mg/kg 100 %
1 females treated at 300 mg/kg 17 %
One female treated at 300 mg/kg had to be killed in extremis 3 hours post-dose and the three females treated at 2000 mg/kg died spontaneously within the first hour after treatment.

Clinical signs:

other: The first group of females treated at 300 mg/kg showed a slightly ruffled fur 1 hour after treatment which persisted up to test day 2 in two females. A hunched posture was visible in two females 1-2 hours post-treatment up to the 5- hour observation. A

Gross pathology:

All three females treated at 2000 mg/kg/bw were found with liquid contents in their stomachs. Otherwise, no macroscopic findings were recorded at necropsy.

Other findings:

no data

Applicant's summary and conclusion

Interpretation of results:

Category 4 based on GHS criteria

Remarks:

Migrated information: H302, harmful if swallowed

Conclusions:

The median lethal dose of test item after single oral administration to female rats, observed over a period of 14 days is: 300 mg/kg body weight < LD50 (female rat) < 2000 mg/kg body weight. Based on the proposed read across approach, the LD50 of the substance registered can be assumed to be in the same range as for the analogue substance.

Executive summary:

The acute oral toxicity to rats of test item (analogue substance) was being investigated following the testing protocol as given in OECD guideline 423.

Three groups, each of three female rats were treated with the test item by oral gavage administration at a dosage of 300 mg/kg (two groups) or 2000 mg/kg (one group) body weight. The test item was diluted in vehicle (purified water) at a concentration of 0.2 g/mL or 0.03 g/mL and administered at a dosing volume of 10 mL/kg.

The animals were examined daily during the acclimatization period and mortality, viability and clinical signs were recorded. All animals were examined for clinical signs within the first 30 minutes and approximately 1, 2, 3 and 5 hours after treatment on day 1 and once daily during test days 2-15. Mortality/viability was recorded at approximately 30 minutes, 1, 2, 3 and 5 hours after administration on test day 1 (with the clinical signs) and twice daily during days 2-15. Body weights were recorded on day 1 (prior to administration) and on days 8 and 15.All animals were necropsied and examined macroscopically.

The following animals were treated and percentage of mortality was observed:

3 females treated at 2000 mg/kg                  100 %

1 females treated at 300 mg/kg                    17 %

One female treated at 300 mg/kg had to be killed in extremis 3 hours post-dose and the three females treated at 2000 mg/kg died spontaneously within the first hour after treatment.

The median lethal dose of test item after single oral administration to female rats, observed over a period of 14 days is: 300 mg/kg body weight < LD50 (female rat) < 2000 mg/kg body weight.

Given the applicability of the proposed read across approach (see IUCLID chapter 13), the respective LD50 values of the substance registered can safely be assumed to be in the same range as for the analogue substance, as (1) the molecules of the substance registered and the analogue substance show to a large extent structural similarities, and (2) furthermore the molecular weight of the analogue substance and the substance registered only differ by 2.5%.