Chromium iron oxide

Endpoint summary

Description of key information

Acute oral toxicity: LD50 > 5000 mg/kg bw (OECD 423 (2001); GLP compliant; female rats)

Acute toxicity, inhalation: LC50 >5.01 mg/L air.  MMAD: 3.656 µm (GSD: 3.08)

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:

From 2021-09-27 to 2021-10-14

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2001-12-17

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

signed 2019-11-29.

Test type:

acute toxic class method

Limit test:

yes

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature and keep dry in closed containers.

Species:

rat

Strain:

Wistar

Remarks:

Crl: WI(Han)

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 8 - 9 weeks old
- Weight at study initiation: 162 - 163 g
- Fasting period before study: 18 hours
- Housing: animals were kept in groups of three animals in IVC cages, type III H, polysulphone cages on Altromin saw fibre bedding.
- Diet (ad libitum): Altromin 1324 maintenance diet for rats and mice (manufacturer: Altromin Spezialfutter GmbH & Co. KG, Lage, Germany)
- Water (ad libitum): tap water, sulphur acidified to a pH value of approximately 2.8.
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Air changes (per hr): 10 times
- Photoperiod (hrs dark / hrs light): 12 / 12

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

sterile

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 2.5 g/5 mL
- Justification for choice of vehicle: based on the outcome of the solubility test. The vehicle was chosen due to its non-toxic characteristics.
- Lot no.: 2005066

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

DOSAGE PREPARATION:
The test item was suspended with the vehicle. Homogeneity of the test item in the vehicle was maintained by vortexing the prepared suspension thoroughly before each dose administration.

CLASS METHOD:
- Rationale for the selection of the starting dose: a dose of 5000 mg/kg bw has been selected for the following reasons: the study will be used for EU countries implementing the CLP regulation as well as for non-EU countries implementing the GHS regulation. Furthermore, the test item shows a low water solubility. As the bioavailability (and thus solubility) of the test item is a key determinant of toxicity, low toxicity is expected, therefore justifying an initial testing at the limit dose of 5000 mg/kg bw.

Doses:

5000 mg/kg bw

No. of animals per sex per dose:

3 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: a careful clinical examination was made several times on the day of dosing (at least once during the first 30 minutes and with special attention given during the first 4 hours post-dose). Thereafter, the animals were observed for clinical signs once daily. Observations on mortality/morbidity were made at least once daily.
Animals were weighed on day 1 (prior to the administration) and on days 8 and 15.
- Necropsy of survivors performed: yes, at the end of the observation period the animals were sacrificed and all animals were subjected to gross necropsy and examined macroscopically for gross pathological changes. In the absence of gross pathological changes no tissues were preserved for a possible histopathological evaluation.

Statistics:

No statistical analysis could be performed (the method used is not intended to allow a calculation of a precise LD50 value).

Preliminary study:

No

Sex:

female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality was observed during the study.

Clinical signs:

other: The test item showed no acute oral toxicity characteristics after a single dose administration at a dosage of 5000 mg/kg bw. Piloerection was observed on day 1 in all animals but no specific findings were observed thereafter (from day 2 to 15).

Body weight:

other body weight observations

Remarks:

None of the animals showed weight loss during the observation period. Throughout the 14-day observation period, the weight gain of the animals was within the normal range of variation for this strain.

Gross pathology:

No specific gross pathological changes were recorded for any animal.

Interpretation of results:

GHS criteria not met

Conclusions:

LD50 (female rats) > 5000 mg/kg bw
According to the Regulation (EC) No 1272/2008 and subsequent adaptations, the substance is not acutely toxic via the oral route.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The study fulfils the rquirements for acute oral toxicity under REACH (Regulation (EC) 1907/2006).

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:

2010-04-07 to 2010-05-24

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Version / remarks:

2009-09-07

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

signed 2009-11-12

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research models and Services, Germany GmbH, Sandhofer Weg 7, 97633 Sulzfeld, Germany
- Age at study initiation: males: 49 days; females: 63 days
- Weight at study initiation: males: 222- 228 g; females: 206 - 222 g
- Fasting period before study: Feeding was discontinued approx. 16 hours before exposure.
- Housing: granulated textured wood (Granulat A2, J. Brandenburg, 49424 Goldenstedt, Germany) was used as bedding material for the cages. During the 14-day observation period the animals are kept by sex in groups of 2-3 animals in MAKROLON cages (type III plus).
- Diet: Commercial diet, ssniff® R/M-H V1534 served as food (ssniff Spezialdiäten GmbH, 59494 Soest, Germany).
- Water (ad libitum): Drinking water
- Acclimation period: At least 5 days; The animals were acclimatised to the test apparatus for approx. 1 hour on 2 days prior to testing, The restraining tubes did not impose undue physical, thermal or immobilization stress on the animals.

ENVIRONMENTAL CONDITIONS
- Temperature: 22 °C +/- 3 °C (maximum range)
- Relative humidity: 55 % +/- 15 % (maximum range)
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: dust

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: The study was carried out using a dynamic inhalation apparatus (RHEMA_LABORTECHNIK, 65719 Hofheim/Taunus, Germany) (air changes/h (≥ 12 times)) with a nose-only exposure of the animals according to KIMMERLE & TEPPER. The apparatus consists of a cylindrical exposure chamber (volume 40 L) which holds the animals in pyrex tubes at the edge of the chamber in a radial position.

- System of generating particulates/aerosols: The dust of the test material was generated with a rotating brush dust generator (RBG 1000, PALAS GmbH partikel und Lasermesstechnik, 76229 Karlsruhe, Germany).
The generator was fed with compressed air (5.0 bar) from a compressor (ALUP Kompressorenfabrik, 73257 Köngen, Germany) (air was taken from the surrounding atmosphere of the laboratory room and filtered using an in-line disposable gas-filter).
At the bottom of the exposure chamber, the air was sucked off at a lower flow rate than it was created by the dust generator in order to produce a homogenous distribution and a positive pressure in the exposure chamber (inflow 900 L/h, outflow 800 L/h).

Before initiating the study with the animals, a pre-test was carried out with the exposure system in order to verify that under the experimental settings chosen, the limit concentration of 5 mg/L air could be achieved by gravimetric analysis.

- Method of particle size determination: An analysis of the particulate size distribution was carried out twice during the exposure period using a cascade impactor according to MAY (MAY, K.R. Aerosol impaction jets, J.Aerosol Sci. 6, 403 (1975), RESEARCH ENGINEERS Ltd., London N1 5RD, UK) .
The dust from the exposure chamber was drawn through the cascade impactor for 5 minutes at a constant flow rate of 5 L/min. The slides were removed from the impactor and weighed on an analytical balance (SARTORIUS, type 1601 004, precision 0.1 mg). Delta of slides’ weight were determined.
The correct functioning of the dynamic separation of particles was controlled microscopically during spot-checks.
The MMAD was estimated by means of non-linear regression analysis. The 32 µm particle size range and the filter (particle size range < 0.5 µm) were not included in the determination of the MMAD in order not to give undue weight to these values.
The GSD of the MMAD was calculated from the quotient of the 84.1%- and the 50%-mass fractions, both obtained from the above mentioned non-linear regression analysis.

- Treatment of exhaust air: The exhaust air was drawn through gas wash-bottles.

- Temperature, humidity, oxygen, carbon dioxide, air flow:: A manometer and an air-flow meter (ROTA Yokogawa GmbH & Co. KG, 79664 Wehr/Baden, Germany) were used to control the constant supply of compressed air and the exhaust, respectively. Flow rates were checked hourly and corrected if necessary.
The oxygen content in the inhalation chamber was 21%. It was determined at the beginning and at the end of the exposure with a DRÄGER Oxygen-analysis test set (DRÄGER Tube Oxygen 67 28 081). Carbon dioxide concentration did not exceed 1%.
Temperature (19.7°C ± 0.3°C) and humidity (52.0% ± 0.3%) were measured once every hour with a climate control monitor (testo 175-HZ data logger).

TEST ATMOSPHERE - The inhalation chamber was equilibrated for at least 15 minutes (t95 approximately 8 minutes).
- Brief description of analytical method used: The actual dust concentration in the inhalation chamber was measured gravimetrically with an air sample filter (Minisart SM 17598 0.45 µm) and pump (Vacuubrand, MZ 2C (Membrane Pump,Vacuubrand GmbH + Co. KG, 97877 Wertheim/Main, Germany)) controlled by a rotameter. Dust samples were taken once every hour during the exposure. For that purpose, a probe was placed close to the animals' noses and air was drawn through the air sample filter at a constant flow of air of 5 L/min for 1 minute. The filters were weighed before and after sampling (accuracy 0.1 mg). The correct loading of the filter was checked by the airflow via the rotameter and by a positive weight increase of the filter after the sampling period of 1 minute.
- Samples taken from breathing zone: yes

TEST ATMOSPHERE (if not tabulated)
- MMAD: 3.656 µm (GSD: 3.08)

Analytical verification of test atmosphere concentrations:

yes

Remarks:

see above ("details on inhalation exposure")

Duration of exposure:

4 h

Remarks on duration:

Exposition started by locating the animals’ noses into the exposure chamber after equilibration of the chamber concentration for at least 15 minutes.

Concentrations:

5.01±0.10 mg Colorante Negro/L air (actual concentration)
5.0 mg/l air (nominal concentration)

No. of animals per sex per dose:

3 males / 3 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: During and following exposure, observations were made. Careful clinical examinations were made at least twice daily until all symptoms subsided, thereafter each working day. Observations on mortality were made at least once daily (in the morning starting on test day 2). Individual weights of animals were determined once during the acclimatisation period, before and after the exposure on test day 1, on test days 3, 8 and 15.
- Necropsy of survivors performed: Yes
Necropsy of all animals was carried out and all gross pathological changes were recorded.
No microscopic examination was carried out as no pathological findings were noted at necropsy.
- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other: Cageside observations included, but were not limited to: changes in the skin and fur, eyes, mucous membranes, respiratory, circulatory, autonomic and central nervous system, as well as somatomotor activity and behaviour pattern.
Particular attention was directed to observation of tremor, convulsions, salivation, diarrhoea, lethargy, sleep and coma. The animals were also observed for possible indications of respiratory irritation such as dyspnoea, rhinitis etc.
Changes in weight were calculated and recorded when survival exceeded one day.

Statistics:

Since no mortality occurred, the calculation of an LC50 was not required.

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.01 mg/L air (analytical)

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: Standard deviation: 0.10 mg/L air; The LC50 cut-off value is 'unclassified'.

Mortality:

No mortality occurred.

Clinical signs:

other: A 4-hour inhalation exposure to Colorante Negro at a concentration of 5.01 mg/L air revealed slight ataxia (0 - 60 minutes), slight to moderate tremor (0 minutes - 3 hours) and slight dyspnoea (0 minutes - 3 hours) in all 3 male and 3 female rats.

Body weight:

All animals gained the expected body weight throughout the study period.

Gross pathology:

No pathological findings were made.

Other findings:

No data

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this study, the 4-hour inhalation LC50 of Colorante Negro is >5.01 mg/L air, and hence, the LC50 cut-off value 'unclassified'.
According to the EC Regulation 1272/2008 and subsequent regulations, the test material is not classified for acute inhalation toxicity.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Acute oral toxicity

One reliable study described in Boldt (2021; OECD 423 (2001); GLP compliant) is considered to be reliable without restrictions and is used as key study for this endpoint. The LD50 was determined to be greater than 5000 mg/kg bw.

 

For acute inhalation toxicity there is one animal study which has been performed according to OECD TG 436 and which shows no signs of acute toxicity after inhalation exposure to chromium iron oxide, indicating a LC50 > 5.01 mg/L.

The 4-hour inhalation exposure to Colorante Negro at a concentration of 5.01 mg/L air revealed slight ataxia (0 - 60 minutes), slight to moderate tremor (0 minutes - 3 hours) and slight dyspnoea (0 minutes - 3 hours) in all 3 male and 3 female rats. No mortality occurred.

 

There are no reliable reports whatsoever on acute dermal toxicity in the public domain. However, the conduct of an acute dermal toxicity study is unjustified as inhalation of the substance is considered as major route of exposure and physicochemical properties of the substance do not suggest a significant rate of absorption through the skin (cf. Annex VIII section 8.5 Column 2 of regulation (EC) 1907/2006). This is underlined by the very low solubility and bioavailability (Pardo Martinez, 2010) of the elements out of this pigment, which can be considered as inert.

Justification for classification or non-classification

Acute oral toxicity

The substance is not acutely toxic via the oral route based on an acute oral toxicity test (OECD 423) and does not require classification according to Regulation (EC) No 1272/2008 and subsequent adaptations.

 

Specific target organ toxicant (STOT) - single exposure: oral

The classification criteria according to regulation (EC) 1272/2008 as specific target organ toxicant (STOT) – single exposure, oral are not met since no reversible or irreversible adverse health effects were observed immediately or delayed after exposure and no effects were observed at the guidance value, oral for a Category 1 classification (C ≤ 300 mg/kg bw) and at the guidance value, oral for a Category 2 classification (2000 mg/kg bw ≥ C > 300 mg/kg bw). No classification required.

 

Acute inhalation toxicity:

The reference Haferkorn, J. is considered as key study for the endpoint acute inhalation toxicity and will be used for classification.

Under the conditions of this study, the 4-hour inhalation LC50 of Colorante Negro is >5.01 mg/L air. According to the EC Regulation 1272/2008 and subsequent regulations, the test material is not classified for acute inhalation toxicity. According to the EC-Commission directive 67/548/EEC and subsequent regulations, the test material is not classified for acute inhalation toxicity.

 

Specific target organ toxicant (STOT) – single exposure: inhalation

The classification criteria acc. to regulation (EC) 1272/2008 as specific target organ toxicant (STOT) – single exposure, inhalation dust/mist/fume are not met since no reversible or irreversible adverse health effects were observed immediately or delayed after exposure and no effects were observed at the guidance value, inhalation dust/mist/fume for a Category 1 classification of 1.0 mg/L/4h and at the guidance value, inhalation dust/mist/fume for a Category 2 classification of 5.0 mg/L/4h. Therefore, no classification is required.

Finally, any category 3 classification should primarily be based on human data. However, such classification is also not warranted, since observations on respiratory irritation in test animals (rats) were not observed.