Reaction product of 3,9-dibromobenzanthrone condensed with 2 equivalents of 1-aminoanthraquinone, subsequently further condensed under oxidative conditions

Endpoint summary

Description of key information

Acute toxicity via oral route

LC50 >2000 mg/kg bw in Wistar rats (OECD 401; EU B.1).

Acute toxicity via inhalation route

LC50: >15,750 mg/m³air

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 October 1993 to 27 October 1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 (Acute Toxicity (Oral))

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS- Source: Hoechst AG
- Age at study initiation: 7 to 8 weeks
- Weight at study initiation: males: 183 to 194 g; females: 174 to 185 g
- Fasting period before study: 16 hours prior to 3-4 hours after dosing
- Housing: 5 per cage- Diet: Altromin 1324 ad libitum
- Water: tap water in plastic bottles ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25°C
- Humidity (%): 35 to 75- Photoperiod (hrs dark / hrs light): 12/12
IN-LIFE DATES: From: 07. Oct To: 21. Oct 1992

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

deionised

Details on oral exposure:

VEHICLE- Deionised water- Justification for choice of vehicle: highly soluble in water

Doses:

2000 mg/kg

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
- Clinical signs: twice daily - on weekends or public holidays: once daily
- Body weight: weekly- Necropsy of survivors performed: yes
The prepared test substance was administered by gavage to fasted animals at the stated dosage. The observation period following treatment lasted for 14 days. Symptoms were recorded twice every day (in the morning and in the afternoon), on week-ends and public holidays only once. During this time the animals were weighed weekly. At the end of the observation period the animals were killed, dissected and examined for macroscopically visible changes.

Statistics:

mean and standard deviation of body weight data

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No deaths.

Clinical signs:

other: During the first day of the study dark red discolored bedding and feces were noted.

Gross pathology:

No abnormalities detected.

Other findings:

None reported

Interpretation of results:

GHS criteria not met

Conclusions:

Based on the results obtained in this study, the oral median lethal dose value (LD50) of the test substance for the male and female rat is greater than 2000 mg/kg bw.

Executive summary:

Acute oral toxicity testing of the test substance in the Wistar rat yielded a median lethal dose (LD50) above 2000 mg/kg bw in both male and female rats.

No signs of intoxication were observed after the application of 2000 mg/kg bw Reaktiv-Rot F-67787. No deaths occurred.

During the first day of the study dark red discolored feces and bedding were noted.

Development of body weight was not impaired.

The animals killed at the end of the observation period showed no macroscopically visible changes. The substance is not classified for oral toxicity.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

Klimisch 1 study - study was conducted by a GLP accredited laboratory in accordance with OECD Guideline 401 and EU Method B.1

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not specified

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

An aerosol of the undiluted test material was generated by passing clean, dry air (-40°C dewpoint) through an Ohio Ball-Jet Nebulizer. The resulting air and aerosol mixture was introduced into the exposure chamber.

GLP compliance:

no

Test type:

traditional method

Limit test:

yes

Species:

rat

Strain:

other: Charles River

Sex:

male/female

Details on test animals or test system and environmental conditions:

Young adult rats were employed as test animals. The rats were selected after having been under observation for at least 5 days to insure their general health and suitability for testing.The animals were housed in stainless steel cages and permitted a standard laboratory diet plus water ad libitum, except during inhalation exposure.

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

clean air

Mass median aerodynamic diameter (MMAD):

other: Not specified

Remark on MMAD/GSD:

No data

Details on inhalation exposure:

Test animals were exposed in a specially constructed inhalation chamber. The chamber was designed so that the animals could be introduced into the test atmosphere after the maximum aerosol concentration was established. Each animal was caged separately during exposure to minimize filtration of inspired air by animal fur.An aerosol of the undiluted test material was generated with an Ohio Ball-Jet Nebulizer. The stream of clean, dry air (-40°C dewpoint) was passed through the nebulizer. The resulting air and aerosol mixture was then introduced into the exposure chamber at the top center, dispersed by a baffle plate and exhausted at the bottom of the chamber. Air flow rate through the system was measured with a rotameter connected upstream of the nebulizer. The rotameter was calibrated with a wet-test meter after the exposure was completed. The average nominal aerosol concentration was calculated by dividing the nebulizer weight loss by the total volume of air used during the test.

Analytical verification of test atmosphere concentrations:

no

Duration of exposure:

4 h

Concentrations:

15,750 mg/m3 air

No. of animals per sex per dose:

10 animal (5 male/5 female)

Control animals:

no

Details on study design:

14 day observation period. During the exposure period, observations were made with respect to incidence of mortality and reactions displayed. At the end the exposure period, the rats were returned to their cages for observation.A body weight was determined for each animal prior to inhalation exposure and for each surviving animal at the end of the observation period. The data were recorded as an index to growth.Gross pathologic examinations were scheduled to be conducted upon all animals which might succumb during the test period and upon those sacrificed at the end of the observation period.

Statistics:

Not specified

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 15 750 mg/m³ air (nominal)

Based on:

test mat.

Exp. duration:

4 h

Mortality:

There were no untoward reactions during exposure or the 14-day observation period which followed.

Clinical signs:

other: There were no untoward reactions during exposure or the 14-day observation period which followed.

Body weight:

The average 2-week body weight gains were within the normal limits.

Gross pathology:

Necropsy, performed on all rats at the end of the observation period, did not reveal any gross pathologic alterations.

Other findings:

No further findings reported.

Results

Group No.	Total Number of Animals Male/Female	Nominal Concentration	Mortality Male-Female	Weight Gain Male-Female (grams)
I	5/5	15,750 mg/m^3air	0/5 – 0/5	120-38

 

Interpretation of results:

GHS criteria not met

Conclusions:

LC50 >15,750 mg/m3 air

Executive summary:

Charles River Rats were exposure for 4 hours.

An aerosol of the undiluted test material was generated by passing clean, dry air (-40°C dewpoint) through an Ohio Ball-Jet Nubulizer. The resulting air and aerosol mixture was introduced into the exposure chamber.

 

There were no untoward reactions during exposure or the 14-day observation period which followed. The average 2-week boy weight gains were within the normal limits. Necropsy, performed on all rats at the end of the observation period, did not reveal any gross pathologic alterations.

 

Acute LC50: >15,750 mg/m³air

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LC50

Value:

15 750 mg/m³ air

Quality of whole database:

Klimisch 2 study - well documented study in accordance with acceptable scientific methods

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not specified

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

Acute dermal toxicity study in rabbits. Observations were conducted for 14 days following dermal application.

GLP compliance:

no

Test type:

standard acute method

Limit test:

yes

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

Young adult albino rabbits of the New Zealand strain were used as test animals. All rabbits had been maintained under observation in the laboratory for at least seven days prior to testing. During the pre-test period, the animals were examined with respect to their general health and suitability as test animals. The rabbits were housed individually in suspended, wire-bottomed cages and maintained on a standard laboratory ration. Food and water were offered ad libitum.

Type of coverage:

occlusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

Twenty-four hours prior to the dermal applications, the backs of the rabbits were shaved free of hair with electric clippers. The shaved area on each animal constituted about 30 percent of the total body surface area. The animals were then returned to their cages to await testing on the following day. The 24-hour waiting period allowed recovery of the stratum corneum from the disturbance which accompanied the close-clipping procedure and permitted healing of any microscopic abrasions possibly produced during the process.The test material was applied at the highest reasonable dose level. The test site was covered by wrapping the trunk of the animal with impervious plastic sheeting which was securely taped in place. This plastic wrap insured close contact of the epidermis and test material. To prevent oral ingestion of the test material, each animal was fitted with a light-weight, flexible plastic collar which was worn throughout the observation period.At the end of the test period, the plastic sheeting and all residual test material were removed.

Duration of exposure:

24 hours

Doses:

3000 mg/kg

No. of animals per sex per dose:

4 animals (2 male/2 female)

Control animals:

no

Details on study design:

The test sites were examined for local skin reactions and the animals were returned to their cages. Observations for mortality, local skin reactions, and behavioral abnormalities were continued for a total of 14 days following the skin applications. Initial, 7 and 14-day body weights were recorded. A necropsy examination was conducted on all animals.In the case of significant mortality following the initial study, additional experiments were conducted at lower dose levels in order to obtain data sufficient to determine the acute dermal median lethal dose (LD50).

Statistics:

At the end of the observation period, the acute dermal median lethal dose (LD50) of the test material was calculated, if possible, using the techniques of Weil, Thompson, and Thompson and Weil.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 3 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality reported

Clinical signs:

other: No pharmacotoxic symptoms were exhibited by any of the rabbits following dermal exposure to CIBANONE Olive S Paste.The test material was moderately to severely irritating to the skin of the albino rabbit. Skin changes at 24 hours were characterized by red

Gross pathology:

Necropsy examination revealed red lungs in rabbit No. 1-M.No other gross pathologic alterations were noted except for the local skin changes as previously described.

Other findings:

None reported

Mortality and Body Weight Data

Dose Level (mg/kg)	Animal Number and Sex	Individual Body Weights (kg)			Number Dead	Percent Dead
		Test Day Number:
		0	7	14	Number Tested
3,000	1-M* 2-M 3-F* 4-F	2.78 2.66 2.90 2.92	2.86 2.72 2.96 3.00	2.90 2.82 3.12 3.10	0/4	0

* The skin at the site of application was abraded

Interpretation of results:

GHS criteria not met

Conclusions:

Acute Dermal LD50: > 3,000 mg/kg

Executive summary:

Young adult albino rabbits of the New Zealand strain were used as test animals.

 

Twenty-four hours prior to the dermal applications, the backs of the rabbits were shaved free of hair with electric clippers. The shaved area on each animal constituted about 30 percent of the total body surface area.

 

The test material was applied at the highest reasonable dose level. The test site was covered by wrapping the trunk of the animal with impervious plastic sheeting which was securely taped in place. This plastic wrap insured close contact of the epidermis and test material.

 

The test material remained in contact with the skin for 24 hours. At the end of this period, the plastic sheeting and all residual test material were removed. The test sites were examined for local skin reactions and the animals were returned to their cages. Observations for mortality, local skin reactions, and behavioral abnormalities were continued for a total of 14 days following the skin applications. Initial, 7 and 14-day body weights were recorded. A necropsy examination was conducted on all animals.

 

No pharmacotoxic symptoms were exhibited by any of the rabbits following dermal exposure to CIBANONE Olive S Paste.

The test material was moderately to severely irritating to the skin of the albino rabbit. Skin changes at 24 hours were characterized by red, well-defined erythema, moderate edema and superficial second degree burns. Mild desquamation was observed at 7 and 14 days.

Necropsy examination revealed red lungs in rabbit No. 1-M.

No other gross pathologic alterations were noted except for the local skin changes as previously described.

 

Acute Dermal LD50: > 3,000 mg/kg

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

3 000 mg/kg bw

Quality of whole database:

Klimisch 2 study - well documented study in accordance with acceptable scientific methods

Additional information

Acute oral toxicity

In the key oral toxicity study, a single oral dose of 2000 mg/kg of the test substance was administered to 10 Wistar rats (5 males and 5 females). No signs of intoxication were observed and no deaths occurred. Development of body weight was not impaired. During the first day of the study dark red discoloured faeces and bedding were noted. The animals killed at the end of the observation period showed no macroscopically visible changes. Therefore, the acute oral median lethal dose (LD50) was determined to be > 2000 mg/kg body weight in both male and female rats.

There are also four supporting acute oral toxicity studies available:

In the first study, the test material (containing ca. 40% of the substance) was administered at 3 doses (1350 mg/kg, 4556 mg/kg and 15380 mg/kg) to young albino Sprague-Dawley rats (4 animals per group, 2 male/2 female). No toxic signs were exhibited by the rats dosed at 1350 mg/kg and 4556 mg/kg. In rats dosed at 15380 mg/kg, hypoactivity was observed at 1 hour, ruffed fur at 3 hours, and black faeces at 5 hours. These clinical signs subsided after 3 days, however areas on the fur and tail remained stained throughout the 14-day observation period. Necropsy examination did not reveal any gross pathologic alterations. Therefore, the acute oral median lethal dose (LD50) was determined to be > 15380 mg/kg body weight in both male and female rats.

In the second study, the test material (containing 43.5% of the substance) was administered at 3 doses (4640 mg/kg, 7750 mg/kg and 10000 mg/kg) to Wistar rats (10 animals per dose, 5 males/5 females). Within 2 hours after treatment the rats in all dosage groups showed sedation, dyspnoea, exophthalmos, curved position and ruffled fur. All animals had recovered within 7 to 8 days; they were killed and autopsied after an observation period of 14 days. On autopsy, no substance related gross organ changes were seen. Therefore, the acute oral median lethal dose (LD50) was determined to be > 10000 mg/kg body weight in both male and female rats.

In the third study, the test material (containing 43.5% of the substance) was administered at 3 doses (4640 mg/kg, 7750 mg/kg and 10000 mg/kg) to Wistar rats (10 animals per dose, 5 males/5 females). Within 2 hours after treatment the rats in all dosage groups showed sedation, dyspnoea, exophthalmos, curved position and ruffled fur. All animals had recovered within 7 to 8 days; they were killed and autopsied after an observation period of 14 days. On autopsy, no substance related gross organ changes were seen. Therefore, the acute oral median lethal dose (LD50) was determined to be > 10000 mg/kg body weight in both male and female rats.

In the fourth study, the test material (containing 43.5% of the substance) was administered at 4 doses (1000 mg/kg, 3000 mg/kg, 10000 mg/kg and 15000 mg/kg) to Wistar rats (10 animals per dose, 5 male/5 female). Reduction in spontaneous motility was noted in the two higher dose groups, however, after 24 hours no clinical signs were reported. No deaths and no gross organ changes were seen. Therefore, the acute oral median lethal dose (LD50) was determined to be > 15000 mg/kg body weight in both male and female rats.

Acute dermal toxicity

Testing by the dermal route does not need to be conducted because the substance does not meet the criteria for classification as acute toxicity or STOT SE by the oral route.

In an acute dermal toxicity study, the test material (containing ca. 40% of the test substance) was administered at a dose of 3000 mg/kg to young, adult, albino rabbits of the New Zealand strain (4 animals, 2 male/2 female). The test material was applied unchanged to the shaved skin of the rabbits for 24 hours; each test site was covered by wrapping the trunk of the animal with impervious plastic sheeting, which was securely taped in place. No toxic symptoms were exhibited by any of the rabbits. The test substance was moderately to severely irritating to the skin of the albino rabbit. Skin changes at 24 hours were characterized by red, well-defined erythema, moderate oedema and superficial second degree burns. Mild desquamation was observed at 7 and 14 days. Necropsy examination revealed red lungs in one male rabbit; no other gross pathologic alterations were noted. Therefore, the acute dermal median lethal dose (LD50) was determined to be > 3000 mg/kg body weight in both male and female rats.

Acute inhalation toxicity

The test substance has a predicted very low vapour pressure and a high melting point and is hence not volatile. Test item synthesis and spray drying is performed in a closed process and the dedusting agents are mixed directly with the wet press cake. Consequently, the final product consists of non-dusty granules or well de-dusted powders and the potential for the generation of inhalable forms is low. In addition the use of this substance will not result in aerosols, particles or droplets of an inhalable size, so exposure to humans via the inhalatory route is unlikely to occur.

In an acute inhalation toxicity study, a single dose of 15750 mg/m3 air of the test material (containing ca. 40% of the test substance) was administered to Charles River rats (10 animals, 5 male/5 female). An aerosol of the undiluted test material was generated by passing clean, dry air (-40°C dew point) through an Ohio Ball-Jet Nebulizer. The resulting air and aerosol mixture was introduced into the exposure chamber; rats were exposed for 4 hours. There were no untoward reactions during exposure or during the 14-day observation period that followed. The average 2-week body weight gains were within the normal limits. Necropsy, performed on all rats at the end of the observation period, did not reveal any gross pathologic alterations. Therefore, the acute inhalation median lethal concentration (LC50) was determined to be 15750 mg/m3 in both male and female rats.

 

Justification for classification or non-classification

The results of these studies do not trigger classification for acute toxicity by the oral, dermal or inhalation routes in accordance with the CLP Regulation.