2,9-dimethylanthra[2,1,9-def:6,5,10-d'e'f']diisoquinoline-1,3,8,10(2H,9H)-tetrone

Endpoint summary

Description of key information

Oral: LD50 (m/f) > 5000 mg/kg bw, rat, according to OECD TG 401, no GLP

Inhalation: Read-across, LC50 (m/f) > 5.2 mg/L air, rat, according to OECD TG 403, GLP compliant

Dermal: LD50 (m/f) > 2500 mg/kg bw, rat, similar to OECD TG 402, no GLP

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:

June 22nd, 1983 - July 6th, 1983

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Qualifier:

according to guideline

Guideline:

other: EEC Directive 79-83 1, Annex V, Part B: Methods For The Determination Of Toxicity 4.1. 1 Acute Toxicity Orally

GLP compliance:

no

Remarks:

but QAU statement included

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

- Physical state: solid (powder)
- Purity: technical grade
- Storage condition of test material: in the dark at room temperature

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: HOECHST AG, Kastengrund, SPF-Zucht
- Age at study initiation: 8 - 10 weeks
- Weight at study initiation: 177 g (males) and 194 g (females)
- Fasting period before study: 16 hours prior and 2 hours after application
- Housing: Makroloncages (Type 4) on softwood granules, in groups of 5
- Diet (e.g. ad libitum): Altromin 1324 (Altromin-GmbH, Lage/Lippe), ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 10
- Air changes (per hr): fully airconditioned rooms
- Photoperiod (hrs dark / hrs light): 12 / 12

Route of administration:

oral: gavage

Vehicle:

other: sesame oil

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 25% (w/v)
- Amount of vehicle (if gavage): 20 ml/kg bw

Doses:

5000 mg/kg bw

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: weekly weighing
- Necropsy of survivors performed: yes

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Remarks on result:

not determinable due to absence of adverse toxic effects

Mortality:

None of the animals died during the observation period.

Clinical signs:

other: The feaces were colored dark brown from 2 hours after application until day 1 post-application.

Gross pathology:

No abnormal findings.

Interpretation of results:

GHS criteria not met

Conclusions:

The LD50 was therefore determined to be greater than 5000 mg/kg body weight.

Executive summary:

In an oral toxicity study according to OECD guideline 401, Wistar rats (5/sex) were treated with the teat substance at 5000 mg/kg bw by single dose (gavage) followed by a 14-day observation period (Hoechst, 1983). None of the animals died during the exposure period. No abnormal clinical observations were observed; the test substance was excreted via the feces (stained feces observed). A normal weight gain was recorded and no abnormal findings were made at necropsy. 

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

5 000 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: inhalation

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Justification for type of information:

see attached justification.

Reason / purpose for cross-reference:

read-across source

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.2 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

not determinable due to absence of adverse toxic effects

Reference 2

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04 Oct 2005 - 07 Nov 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1300 (Acute inhalation toxicity)

Qualifier:

according to guideline

Guideline:

other: EU Guideline 92/69/EEC and 93/21/EEC

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Specific details on test material used for the study:

- Physical state: Solid, black powder
- Analytical purity: > 99%
- Storage condition of test material: Room temperature
- Homogeneous

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Laboratory Animal Services; Wölfersraße 4, 4414 Füllinsdorf, Switzerland
- Age at study initiation: 9 weeks for males and approx. 11 weeks for females.
- Weight at study initiation: males: 260,5 g (mean); females: 205,3 g (mean)
- Housing: singly in cages type DK III (Becker, Germany) without bedding
- Diet: KLIBA mouse / rat laboratory diet 10 mm pellets "GLP", Provimi Kliba SA, Kaiseraugst, Basel Switzerland, ad libitum
- Water: drinking water ad libitum
- Acclimation period: at least 1 week in which they were adapted to the surroundings

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24°C
- Humidity (%): 30 - 70%
- Photoperiod (hrs dark / hrs light): dark cycle of 12 hours

Route of administration:

other: dust aerosol

Type of inhalation exposure:

nose/head only

Vehicle:

other: 2% (w/w) of Aerosil 200

Mass median aerodynamic diameter (MMAD):

>= 3 - <= 3.1 µm

Geometric standard deviation (GSD):

>= 3.1 - <= 3.3

Details on inhalation exposure:

GENERATION OF THE INHALATION ATMOSPHERE
- The test substance was stirred in its container before a sample for dust generation was taken. The test substance was desagglomerated in a mixer under addition of 2% (w/w) of Aerosil 200 before introduction into the dust generator, in order to improve dust aerosol formation. The dust was produced inside the inhalation system with a dosing-wheel dust generator and compressed air.

EXPOSURE
- The exposure system was located inside an exhaust cabin in an air-conditioned laboratory. A supply air flow (compressed air) of 1.5 m³/h was used for the exposure. The exhaust airflow was set to 1.35 m³/h. An air change of about 27 times per hour can be calculated by dividing the supply air flow by the volume of the inhalation system.
 
- The lower amount of exhaust air, which was adjusted by means of a separate exhaust air system, achieved a positive pressure inside the exposure system . This ensured that the mixture of test substance and air was not diluted with laboratory air in the breathing zones of the animals. The animals were exposed to the inhalation atmosphere for 4 hours plus equilibration time of the inhalation systems (t99 about 10 min).

Measurement of operation conditions
- The flows of supply and exhaust air were adjusted and continuously measured with flowmeters. They were recorded four times in about 1-hour intervals. The temperature in the inhalation system was measured continuously with a digital thermometer and recorded four times in about 1-hour intervals. The humidity in the inhalation system was measured with a dielectric probe four times in about 1-hour intervals.
 
- No surveillance of the oxygen content in the inhalation system was performed. The air change was judged to be sufficient to prevent oxygen depletion by the breathing of the animals, and the concentration of the test substance used could not have a substantial influence on oxygen partial pressure.
 
Gravimetric determination of the inhalation atmosphere concentration:
Equipment: balance Mettler AT 250. Preweighed filters were placed into the filtration equipment. By means of the vacuum pump metered volumes of the dust were drawn through the filter. For each sample the dust aerosol concentration in mg/I was calculated from the difference between the preweight of the filter and the weight of the filter after sampling, with reference to the sample volume of the inhalation atmospheres. Mean and standard deviation were calculated for the concentration from the results of the individual measurements. The mean concentration was corrected for the amount of additive used.
 
Particle size analysis:
Definitions
• EACD 50% (effective aerodynamic cutoff diameter 50%) defines the separation characteristic of each impactor stage. 50% of particles with the EACD given are deposited in the pertinent impactor stage; the remainder has reached one of the following stages.
• MMAD (mass median aerodynamic diameter) is the calculated aerodynamic diameter which divides the size distribution in half when measured by mass.
• Geometrical standard deviation (GSD) is the ratio of the estimated 84 percentile to the 50 percentile and indicates the slope of the cumulative particle size distribution curve.
• Inspirable aerosol
Inspirable aerosols can enter the respiratory system. Aerosols with MMAD <100 Nm are considered inspirable (MAK Commission of the DFG (German Research Society, MAK list 1997).
• Respirable aerosol
Respirable aerosols can enter the alveolar region of the lung. Aerosols with MMAD < 4 Nm are considered respirable (MAK Commission of the DFG (German Research Society, MAK list 1997).

Before sampling, the impactor was assembled with preweighed glass-fiber collecting discs, and a backup particle filter. The impactor was connected to the vacuum pump and two samples were taken from the breathing zone of the animals starting not earlier than 30 minutes after the beginning of the exposure. The sample volume was 6 L. After sampling the impactor was taken apart. The collecting discs and the backup particle filter were re-weighed. The amounts of material adsorbed to the walls of the impactor and in the sampling probe (wall losses) were also determined quantitatively. The results from the particle size analysis were not corrected for the additive.

- Exposure conditions:
Supply air (m³/h): 1.5
Exhaust air (m³/h): 1.35
Substance flow (g/h): 36.1
Temperature (°C): 21.8±0.3
Relative humidity (%): 47.7±2.2

- Analytical concentration:
Mean mg/l: 5.2
Standard deviation: 1.0
Nominal concentration mg/l: 24.1

- Particle size analysis:
MMAD (µm): 3.1 and 3.0
Geometrical standard deviation: 3.1 and 3.3

Analytical verification of test atmosphere concentrations:

yes

Remarks:

Gravimetric determination

Duration of exposure:

4 h

Concentrations:

5.2 mg/l (nominal concentration: 24.1 mg/l)

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: A check for overt clinical signs of toxicity or mortality as well as a check for the presence of feed and drinking water was made twice a day on workdays and once daily on weekends and public holidays. Detailed clinical observations were recorded for each animal separately several times during exposure and at least once on each workday of the observation period.
- Body weights: The body weight of the animals was determined just prior to exposure (day 0), weekly thereafter and at the end of the observation period.
- Necropsy of survivors performed: yes. At the end of the observation period the animals were sacrificed with C02 and were subjected to gross-pathological examination

Statistics:

A binomial test was used for statistical evaluation.

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.2 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

not determinable due to absence of adverse toxic effects

Mortality:

No lethality occurred at the tested concentration of 5.2 mg/I during the study period of 14 days.

Clinical signs:

other: Accelerated respiration, pulmonary respiration e sounds, squatting posture, piloerection and smeared fur. Findings were observed from hour 0 of exposure until including study day 3.

Body weight:

The mean body weights of the animals increased throughout the study period.

Gross pathology:

No pathological abnormalities of the organs were observed in all animals at termination of the study.

Other findings:

Contaminated fur was noted during necropsy in all animals.

Analytical concentration:

Mean (mg/l)	standard deviation	nominal concentration (mg/l)
5.2	1	24.1

Particle Size Analysis:

Sample	MMAD (µm)	geometrical standard deviation
1	3.1	3.3
2	3	3.1

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this study the LC50 for male and female rats after dust inhalation was determined to exceed 5 .2 mg/I.

Executive summary:

For determination of the acute inhalation toxicity (single 4-hour-exposure) of the test substance as dust, a GLP compliant study in male and female Wistar rats was performed according to OECD-Guideline method 403, as well as EEC and EPA guidelines. A measured concentration of 5 .2 mg/I was tested (Limit test). Cascade impactor measurements resulted in particle size distributions with mass median aerodynamic diameters (MMADs) of 3.0 and 3.1 µm, which are well within the respirable range. No mortality occurred at the tested concentration. Clinical signs of toxicity comprised visually accelerated respiration, pulmonary respiration sounds, squatting posture, piloerection and smeared fur. Findings were observed from hour 0 of exposure until including study day 3. Moreover, contaminated fur was observed in all animals from day 0 onward until termination of the study. The mean body weights of the animals increased throughout the study period. No pathological abnormalities of the organs were observed in all animals at termination of the study. Contaminated fur was noted during necropsy in all animals. Under the conditions of this study the LC50 for male and female rats after dust inhalation was > 5 .2 mg/l.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating conc.

Value:

5 200 mg/m³ air

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Qualifier:

according to guideline

Guideline:

other: D.N.NOAKES und D.M.SANDERSON (A Method for Determining the Dermal Toxicity of Pesticides; Brit.Journ.Ind.Med. 26, 59, 1969

GLP compliance:

no

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Wiga, SPF-bred
- Weight at study initiation: males: 141 g; females: 132 g

Type of coverage:

not specified

Vehicle:

water

Details on dermal exposure:

TEST SITE
- Area of exposure: back, area of 50 cm²

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 5 ml/kg
- Concentration (if solution): 50% solution

Duration of exposure:

Not specified

Doses:

2500 mg/kg bw

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: daily
- Necropsy of survivors performed: yes

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 500 mg/kg bw

Based on:

test mat.

Remarks on result:

not determinable due to absence of adverse toxic effects

Mortality:

None of the animals died during the observation period.

Clinical signs:

other: No abnormal findings. Red-brown staining at the application site observed.

Gross pathology:

No abnormal findings.

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions chosen, the test substance was non-toxic to the experimental animals.

Executive summary:

The acute dermal toxicity of the test substance was determined in a study similar to OECD TG 402 but without GLP compliance. Five male and five female Sprague-Dawley rats were exposed to a 50 % suspension of the test substance in water. The test substance was administered in a single dose of 2500 mg/kg bw. After exposure the animals were observed for a period of 14 days. No mortality was observed during the test. A red-brown staining by the pigment at the application site could be observed observed. No other abnormal findings (clinical signs, body wight, necopsy) were noted after exposure.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

2 500 mg/kg bw

Additional information

Oral toxicity

In an oral toxicity study according to OECD guideline 401, Wistar rats (5/sex) were treated with the teat substance at 5000 mg/kg bw by single dose (gavage) followed by a 14-day observation period (Hoechst, 1983). None of the animals died during the exposure period. No abnormal clinical observations were observed; the test substance was excreted via the feces (stained feces observed). A normal weight gain was recorded and no abnormal findings were made at necropsy. The LD50 was therefore determined to be greater than 5000 mg/kg body weight.

 

Supporting information:

In a supporting study following a protocol comparable to OECD guideline 401, Sprague-Dawley rats (5/sex) were treated with the test substance at 5000 mg/kg bw by single dose (gavage) followed by a 14-day observation period (BASF, 1980). None of the animals died during the exposure period. No abnormal clinical observations were observed; the test substance was excreted via the feces. A normal weight gain was observed and no abnormal findings were made at necropsy. The LD50 was therefore set at greater than 5000 mg/kg body weight.

In another OECD guideline 401 comparable study, female Sprague-Dawley rats (5/sex) were administered the test article at 2000 mg/kg bw by single dose (gavage) followed by a 14-day observation period (Hazleton Laboratories Europe Ltd., 1978). None of the animals died during the exposure period. No abnormal clinical observations were observed. A normal weight gain was observed and no abnormal findings were made at necropsy. The LD50 observed in this study was greater than 2000 mg/kg body weight.

Two more studies comparable to OECD guideline 401 are available which investigate the acute oral toxicity of the test article or of a mixture containing the test article (both BASF, 1976). Rats (n = 5) were treated with the test material at 10000 mg/kg bw by single dose (gavage) followed by a 7-day observation period. None of the animals died during the exposure period. Clinical signs in each study included apathy and dyspnea, respectively. No abnormal findings were made at necropsy. The LD50 in both studies was greater than 10,000 mg/kg bw for the material tested.

 

Inhalation toxicity

Regarding inhalation toxicity, no key information is available for the test substance. To fill this data gap, a read-across to the category member EC 479-300-2 was performed. For determination of the acute inhalation toxicity (single 4-hour-exposure) of the test substance, a GLP compliant study in male and female Wistar rats was performed according to OECD-Guideline method 403, as well as EEC and EPA guidelines. A measured dust concentration of 5.2 mg/L was tested (Limit test). Cascade impactor measurements resulted in particle size distributions with mass median aerodynamic diameters (MMADs) of 3.0 and 3.1 µm, which are well within the respirable range. No mortality occurred at the tested concentration. Clinical signs of toxicity comprised visually accelerated respiration, pulmonary respiration sounds, squatting posture, piloerection and smeared fur. Findings were observed from hour 0 of exposure until including study day 3. Moreover, contaminated fur was observed in all animals from day 0 onward until termination of the study. The mean body weights of the animals increased throughout the study period. No pathological abnormalities of the organs were observed in all animals at termination of the study. Contaminated fur was noted during necropsy in all animals. Under the conditions of this study the LC50 for male and female rats after dust inhalation was > 5 .2 mg/l.

 

Supporting information:

In an inhalation toxicity study comparable to OECD guideline 403, single maximum concentration of the test material was administered to 20 Sprague-Dawley rats (10 male, 10 female) as dust by inhalation (nose only) over a period of 4 hours. A further 10 rats (5 male, 5 female) were exposed under similar conditions to an atmosphere of filtered air. The concentration of test material, as measured by gravimetric analysis was 0.41 mg/l. (Hazleton, 1978). It is however not verified that the saturated dust concentration is the maximum possible. The mass median aerodynamic diameter (MMAD) was 0.87 µm. No deaths occurred during exposure or during the observation period as a result of exposure to the test material. One male treated rat suffocated as a result of turning around in the restraining tube. A planned sacrifice was made of one female treated rat, one male control and one female control rat after exposure to assess the degree of primary lung irritation. Treated rats had chromodacryorrhoea for a few hours after exposure and their pelts remained stained red throughout the observation period. Control rats had nasal secretion and chromodacryorrhoea on the day of exposure presumably an irritant effect of the restraining tubes. Their pelts were ruffled, a condition which persisted throughout the observation period. The mean body weights of the male and female control rats rose throughout the experiment, slowly until day 2 but thereafter more rapidly. The mean body weights of the treated rats were depressed following exposure but raised steadily throughout the observation period. All the control rats and 6 male and 7 female treated rats, sacrificed after the 14 day observation period showed a moderate degree of pulmonary congestion and edema was present in some control rats. A similar degree of congestion was also noted in the female treated rat examined immediately after exposure, but congestion was more severe in the male, which had suffocated. No congestion was noted in the two control rats sacrificed at the same time, and no other abnormalities were noted in any of the control animals. The LC50 was > 0.41 mg/L.

 

Dermal toxicity

In the key study comparable to OECD guideline 402, 5 Sprague-Dawley rats of each sex were treated with the test substance at 2500 mg/kg bw by single dose followed by a 14-day observation period (BASF AG, 1976). None of the animals died during the exposure period. No abnormal clinical observations were observed and no abnormal findings were reported during necropsy. The LD50 was greater than 2500 mg/kg bw.

In a supporting dermal toxicity study comparable to OECD guideline 402, Sprague-Dawley rats (5/sex) were administered a mixture containing 18.5% of the test substance at 5 ml/kg bw by single dose followed by a 14-day observation period (BASF, 1976). None of the animals died during the exposure period. No abnormal clinical observations were observed and no abnormal findings were reported at necropsy.

 

Further toxicological data of category members:

The test article belongs to the "perylene based organic pigments" category (see attached document for details on category members and for read across justification). According to the category approach, missing toxicity endpoints can be addressed with data available for other category members. Regarding acute toxicity, reliable data are available for the test article and for other members of the "Perylene based pigments category". All of these data are taken into account for the evaluation and assessment of the acute toxicity of the test article.

Additional information is available for the oral and the inhalation route:

In several studies performed with other substances from the Perylene category the potential for oral toxicity was found to be very low. None of these studies raised any concerns regarding acute toxicity after oral application and therefore none of the substances requires classification. The LD50 values observed for these compounds were greater than 5000 mg/kg bw in alle studies performed.

To assess the acute inhalation toxicity, other category members were tested in studies according or similar to OECD 403 guideline. In all studies, rats were exposed to analytically verified dust aerosols for a duration of 4 hours. Except for one study with a single case of mortality all animals survived the procedure. The observed clinical signs included accelerated respiration, pulmonary respiration sounds, squatting posture, piloerection, flight behavior and smeared fur. No pathological abnormalities of the organs were observed at termination in all animals in any of the studies. The LC50 was always greater than tested limit dose respectively.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. No mortality occurred at the limit dose of 2000 mg/kg bw. As a result, the substance is not considered to be classified for acute oral, inhalation or dermal toxicity under Regulation (EC) No. 1272/2008, as amended for the fourteenth time in Regulation (EC) No. 2020/217.