5,5'-(1H-isoindole-1,3(2H)-diylidene)dibarbituric acid

Endpoint summary

Description of key information

key, oral, rat: LD50 > 10000 mg/kg bw; transient signs of toxicity: yellow dyed feces (non-GLP, similar to OECD 401; BASF SE, 1973)
read-across, CAS 76199-85-4, key, inhalative, rat: LC50 >5.42 mg dust/L air; signs of toxicity: no mortality and no toxicity observed (non-GLP, similar to  OECD 403; BASF SE, 1982)
key, dermal, rat: LD50 > 2500mg/kg bw; transient signs of toxicity: no abnormality detected (non-GLP, similar to OECD 402; BASF SE, 1973)
supporting study, intraperitoneal, mouse: LD50 > 6400 mg/kg bw; signs of toxicity: no mortality; accelerated respiration, spastic gait and stretching, hunched body posture, closed eyes (non-GLP, non-guideline, BASF SE, 1973)

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

yes

Remarks:

(application volume exceeds 20 mL/kg bw, 7 day observation period)

GLP compliance:

no

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
-Source: Gassner
- Weight at study initiation: mean: males 214 g; females 186 g

ENVIRONMENTAL CONDITIONS
not reported

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

aqueous solution

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 30 % aqueous suspension in CMC solution

MAXIMUM DOSE VOLUME APPLIED:
33.3 mL/kg bw.

Doses:

8000 and 10000 mg/kg bw

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 7 days
- Frequency of observations and weighing: Weighing was only performed at the beginning of the study for dose calculation. Observation of clinical signs was several times on the day of administration and once daily afterwards with the exception of weekends and on holidays.
- Necropsy of survivors performed: yes

Statistics:

not performed

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 10 000 mg/kg bw

Based on:

test mat.

Mortality:

no mortality observed

Clinical signs:

other: no clinical signs observed

Gross pathology:

no abnormalities observed

Other findings:

yellow coloured faeces

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

10 000 mg/kg bw

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:

03.11.1981 - 17.11.1981

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

GLP compliance:

no

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

Pigment Yellow 185

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: WIGA Versuchstierzuchtanstalt, Sulzfeld, Germany, outbred strain: CAW-ICO-Wiga
- Age at study initiation: about 8 - 10 weeks
- Weight at study initiation: male animais 318 +/- 27 g, female animals 228 +/- 19 g
- Housing: five per cage, cages of Becker, type D III, without bedding.
- Diet: SSNIFF R complete diet for rats and mice, manufacturer: SSNIFF-Versuchstierdiaeten GmbH, Soest, Germany, ad libitum
- Water: tap water ad libitum during the observation period


ENVIRONMENTAL CONDITIONS
- Temperature (°C): temperature 22 +/- 2
- Humidity (%): 55 +/- 5
- Photoperiod (hrs dark / hrs light): 12 h/12 h

Route of administration:

inhalation: dust

Type of inhalation exposure:

nose/head only

Mass median aerodynamic diameter (MMAD):

1.7 µm

Geometric standard deviation (GSD):

4

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Read-nose inhalation system INA 20 (glass-steel construction, BASF AG)
- Exposure chamber volume: 55 L
- Method of holding animals in test chamber: The animals are restrained in tubes and their snouts project into the inhalation chamber.
- Source and rate of air: 1500 L/h compressed air by the injector and 1500 L/h ventilator air as dilution air.
- System of generating particulates/aerosols: A mixture of dust and air was generated by means of dust metering equipment (BASF).
By means of a dust generator the substance to be tested was generated into a dust aerosol, which was passed into the inhalation system.
An automatic vibrator was used for dust generation. The concentration was adjusted by varying the apertural width and varying the amplitude of oscillations of the metering beaker.
- Method of particle size determination: Andersen Stack Sampler Mark III Millipore vacuum compressed air pump XX 60 220 50, limiting orifice 3 L/min, Millipore sampling probe, internal diameter 6.9 mm, 30 minutes after the beginning of the test at the earliest, one sample was taken for the particle size analysis. Before the sampling, the impactor was equipped with glass-fiber collecting discs and a backup particle filter.
The impactor was connected to the pump and the test apparatus, and one sample (9 L) was removed.
The impactor was taken apart, and the collecting discs and the backup particle filter were weighed. The contents of the pre-impactor were determined gravimetrically.
- Pressure in air chamber: 1500 L/h compressed air by the injector and 1500 L/h ventilator air as dilution air
- To ensure, that the mixture of the test substance and air was not diluted with fresh air, the air suction system was reduced about 10 % compared to the supply air system (excess pressure).

TEST ATMOSPHERE
- Brief description of analytical method used:
• Vacuum compressed air pump (Millipore) XX 60 220 50,
• Filtration equipment with probe (Millipore) (internal diameter: 4 mm),
• Filter: MN 85/90 Bf (d = 4.7 cm),
• Balance: Cahn 26
Gravimetric determination of the concentration: the preweighed filter was placed into a filtration equipment. By means of a vaccuum compressed air pump a volume of the dust aerosol characterized by a limiting orifice was drawn through the filter.
The dust concentration in mg/L 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.
- Samples taken from breathing zone: yes,
• Sampling: Velocity: 1.25 m/s, amount: 1 L, probe diameter: 4 mmm, frequency: half-hourly, in the immediate vicinity of the animal noses


- Particle size distribution: EACD 50% (effective aerodynamic cutoff diameter 50%);
Stage: Pre-impactor / EACD 50 % (µm): 26.6
Stage: Cascade impactor: Backup filter / EACD 50 % (µm): <1.2 / Percentage distribution (%): 41.14
Stage: Cascade impactor: 7 / EACD 50 % (µm): 1.2 / Percentage distribution (%): 18.69 / Cumulative distribution (%): 41.14
Stage: Cascade impactor: 5 / EACD 50 % (µm): 2.8 / Percentage distribution (%): 22.28 / Cumulative distribution (%): 59.83
Stage: Cascade impactor: 4 / EACD 50 % (µm): 5.5 / Percentage distribution (%): 8.40 / Cumulative distribution (%): 82.11
Stage: Cascade impactor: 3 / EACD 50 % (µm): 8.5 / Percentage distribution (%): 5.22 / Cumulative distribution (%): 90.51
Stage: Cascade impactor: 1 / EACD 50 % (µm): 18.2 / Percentage distribution (%): 2.14 / Cumulative distribution (%): 95.73
Stage: Cascade impactor: 0 / EACD 50 % (µm): 29.5 / Percentage distribution (%): 2.13 / Cumulative distribution (%): 97.87

A respirable dust fraction of 90.5% was obtained from the results of the particle size analysis.

- MMAD (Mass median aerodynamic diameter) 50 % = 1.7 µm/ GSD (Geometric st. dev.): 4.0

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

5.42 mg/l (analytical), 19.7 mg/l (nominal)

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: daily observation, weighing; before start of application, then on day 7 and day 14,
- Necropsy of survivors performed: yes, with grosspathological examination.
- Other examinations performed: body weight

Statistics:

The statistical evaluation of the concentrations-response relationship was carried out in accordance with the binomial test (Wittig, H.: Mathematische Statistik 1974, pp.32 - 35) using tables of the BASF Computer Center.
The particle size was determined in accordance with mathematical and graphical methods of evaluating particle measurements (Silverman, L.: Particle Size Analysis in Industrial Hygiene, 1971, pp. 235-259).

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.42 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: No mortality and no toxicity observed.

Mortality:

no mortality

Clinical signs:

other: During exposure: no abnormalities; After exposure: fur discolored by the test substance, particularly the region of the head; otherwise no abnormalities.

Body weight:

The body weight of the animals showed no adverse effects in comparison with that of the control.
Mean body weight (male / female)
- Before the study
Treatment group: 318 / 228
Control: 307 / 230
- After 7 days
Treatment group: 345 / 242
Control: 340 / 240
- After 14 days
Treatment group: 368 / 251
Control: 367 / 249

Gross pathology:

No abnormalities detected.

Interpretation of results:

practically nontoxic

Remarks:

Migrated information

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating conc.

Value:

5 420 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:

other: Comparable to guideline study with acceptable restrictions (occlusive treatment, 24 h exposure, incomplete documentation)

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

yes

Remarks:

(occlusive treatment, 24 h treatment)

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, Ottobrunn, Germany
- Weight at study initiation: mean weight: males 151 g, females 129 g

ENVIRONMENTAL CONDITIONS: not reported

Type of coverage:

occlusive

Vehicle:

water

Details on dermal exposure:

TEST SITE
- Area of exposure: dorsal, p.c., 50 cm²
- % coverage: > 10 %

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2.5 g/kg animal
- Concentration (if solution): 50 % aequous solution
- For solids, paste formed: yes

Duration of exposure:

24 h

Doses:

2.5 g/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: No weighing was done. Observation was several times at the day of exposure and daily on working days.
- Necropsy of survivors performed: yes

Statistics:

None

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 500 mg/kg bw

Based on:

test mat.

Remarks on result:

other: No mortality and no clinical signs were observed.

Mortality:

0/10 animals died after the exposure

Clinical signs:

other: no abnormalities detected

Gross pathology:

9/10 no abnormalities detected
1/10 right peak pulmonary lobe with chronic Bronchopneumonia

Other findings:

- Other observations: after 24 h local orange substance residues

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

discriminating dose

Value:

2 500 mg/kg bw

Additional information

There are reliable studies available to assess the acute oral and dermal toxicity of the test substance. The studies were performed in 1973 when characterization of powders in regard to particles with a size of < 100 nm was not routinely performed. Doses applied exceeded the maximum doses later prescribed in OECD testing guidelines. Retrospective analysis did not allow any conclusion on whether the test material was a a nanomaterial or not. However, organic pigments are all powders that often have a nanosize fraction. They consistently show absence of a hazard if tested up to the limit dose in studies with oral and dermal dosing (Stratmann, et al. Indicators for lack of systemic availability of organic pigments, https://doi.org/10.1016/j.yrtph.2020.104719, Regulatory Toxicology and Pharmacology Volume 115, August 2020, 104719). The publication lists 113 and 35 pigments tested for acute oral and dermal toxicity, respectively and in none of the studies, treatment-related morbidities were observed. Therefore, the available experimental data is considered adequate to assess the hazard of the pigment both in the bulk and in the nano form.

For acute inhalative toxicity there is only an acute toxicity study available for for the analogous test substance CAS 76199-85-4. In general support of read-across, all acute studies for PY 185 are listed in the table and provided as robust study summaries.

Oral

A study was performed to assess the acute toxicity following oral administration of the test substance in rats similar to OECD guideline 401 with deviations (BASF SE, 1973). To a group of five animals per sex and per dose a single oral dose of the test material preparation (dose volume: 20 ml/kg bw) in an aqueous solution of carboxymethyl cellulose at a dose level of 8000 and 10000 mg/kg bw was given. The animals were examined for clinical signs several times on the day of administration and once daily afterwards with the exception of weekends and on holidays. The observation period was 7 days instead of 14 days as recommended in OECD 401. Surviving animals were necropsied at the end of the study. No clinical signs were observed during the study. Only yellow colored feces were noticed. All animals survived until the end of the study period. At necropsy no abnormalities were observed.

Under the conditions of this study the median lethal dose of the test substance after oral application was found to be greater than 10000 mg/kg body weight for male and female rats.

Inhalative:

In a study evaluating acute toxicity following inhalative administration Sprague-Dawley rats were exposed to the analogous test substance CAS 76199-85-4 according to OECD guideline 403 without deviations (BASF 1982). Five male and female animals were exposed to 5.42 mg/l (analytical) test substance in a head-nose inhalation system for 4 hours. The MMAD 50% (mass median aerodynamic diameter 50%) was 1.7 µm, calculated from the results of the particle size analysis and the respirable dust fraction was calculated to be 90.5%. During the observation period of 14 days, the animals were examined for clinical signs daily and then necropsied with gross pathological examination at the end of the observation period. No mortality was observed during the course of the study. No clinical signs were observed during the study period. Only the fur was discolored by the test substance, particularly the region of the head.

Under the conditions of this study the median lethal concentration of the test substance after inhalative exposure was found to be greater than 5.42 mg dust/L air for male and female animals.

Dermal

A study was performed to assess the acute toxicity following dermal administration of the test substance in Sprague-Dawley rats similar to OECD guideline 402 with deviations (BASF SE, 1973). Five animals per sex were treated with the test substance at 2500 mg/kg bw by dermal occlusive application. The test item was diluted in vehicle (an aqueous solution of carboxymethyl cellulose) at a concentration of 50%. The application period was 24 hours. During the observation period of 14 days, the animals were examined for clinical signs several times on the day of exposure and daily afterwards on working days. All animals survived and no clinical signs were observed during the study period. In nine of ten animals no abnormalities were detected at necropsy. In only one of ten animals right peak pulmonary lobe with chronic Bronchopneumonia was observed in gross pathology.

Under the conditions of this study the median lethal dose of the test substance after dermal application was found to be greater than 2500 mg/kg body weight for male and female animals.

Intraperitoneal:

A non guideline study was performed to assess the acute toxicity following single intraperitoneal administration of the test substance in mice (BASF 1973). In that supporting study animals were treated with the test substance in the doses 3200 and 6400 mg/kg bw in a 30% aqueous solution with carboxymethyl cellulose as vehicle. During the observation period of 7 days, the animals were examined for clinical signs. Dyspnea and spastic gait were observed. All animals survived until the end of the study period. Therefore, the approximative median lethal dose is > 6400 mg/kg bw. At necropsy intraabdominal deposits of test substance and slightly colored organs were observed.

Under the conditions of this study the approximative median lethal dose of the test substance after single intraperitoneal application was found to be greater than > 6400 mg/kg body weight for male and female animals.

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 or dermal toxicity under Regulation (EC) No. 1272/2008, as amended for the fourteenth time in Regulation (EC) No. 2020/217.