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Description of key information

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:
08 April 2015 - 29 September 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study was performed at a GLP laboratory in accordance with OECD, EU and US testing guidelines for the assessment of Acute Oral Toxicity.
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Deviations:
yes
Remarks:
A deviation in the daily mean humidity was observed during the study, however this deviation did not affect the validity of the study.
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Deviations:
yes
Remarks:
A deviation in the daily mean humidity was observed during the study, however this deviation did not affect the validity of the study.
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Deviations:
yes
Remarks:
A deviation in the daily mean humidity was observed during the study, however this deviation did not affect the validity of the study.
Qualifier:
according to
Guideline:
other: JMAFF Guidelines (2000), including the most recent revisions.
Deviations:
yes
Remarks:
A deviation in the daily mean humidity was observed during the study, however this deviation did not affect the validity of the study.
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Approx. 9-10 weeks old
- Weight at study initiation: Weight variation did not exceed +/- 20% of the sex mean.
- Fasting period before study: Animals were deprived of food overnight prior to dosing and until 3-4 hours after administration of the test substance.
- Housing: 3 animals per cage in labeled Makrolon cages containing sterilized sawdust as bedding material and paper as cage-enrichment.
- Diet (e.g. ad libitum): Free access to pelleted rodent diet
- Water: ad libitum
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24°C
- Humidity (%): 40 to 70%
- Air changes (per hr): 10
- Photoperiod: 12-hour light/12-hour dark cycle
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 20-23%
- Amount of vehicle (if gavage): 10 mL/kg
- Justification for choice of vehicle: The vehicle was selected based on trial preparations and on test substance data supplied by the Sponsor.

MAXIMUM DOSE VOLUME APPLIED: 2000 MG/KG
Doses:
2000 mg/KG
No. of animals per sex per dose:
Each dose group consisted of 3 animals (female).
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were subjected to daily observations and weekly determination of body weight.
- Necropsy of survivors performed: yes, Macroscopic examination was performed after terminal sacrifice (Day 15).
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality occurred.
Clinical signs:
Hunched posture and/or piloerection were noted animals on Day 1. Blue discolouration of the skin/fur, faeces and urine were noted for all animals on Day 1 and/or 2. This was considered to be due to the test substance.
Body weight:
The mean body weight gain shown by the animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain.
Gross pathology:
No abnormalities were found at macroscopic post mortem examination of the animals.
Other findings:
.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test substance does not have to be classified and has no obligatory labelling requirement for acute oral toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).
Executive summary:

The test substance was administered by oral gavage to two subsequent groups of three female Wistar rats at 2000 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice (Day 15).

No mortality occurred.

Hunched posture and/or piloerection were noted animals on Day 1.

Blue discolouration of the skin/fur, faeces and urine were noted for all animals on Day 1 and/or 2. This was considered to be due to the test substance.

The mean body weight gain shown by the animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain.

No abnormalities were found at macroscopic post mortem examination of the animals. The oral LD50 value of K1600 black dye in Wistar rats was established to exceed 2000 mg/kg body weight.

According to the OECD 423 test guideline, the LD50 cut-off value was considered to exceed 5000 mg/kg body weight.

Based on these results, K1600 black dye does not have to be classified and has no obligatory labelling requirement for acute oral toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating dose
Value:
2 000 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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study was performed at a GLP laboratory in accordance with OECD, EU and US testing guidelines for the assessment of Acute Inhalation Toxicity.
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: Rat: Crl:WI(Han) (outbred, SPF-Quality)
Sex:
male/female
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Analytical verification of test atmosphere concentrations:
yes
Remarks:
gravimetric
Duration of exposure:
ca. 4 h
Concentrations:
The time-weighted mean actual concentration was 5.1 ± 0.15 mg/L. The nominal concentration
(amount of test item used divided by the volume of pressurized air used) was 22 mg/L. The generation
efficiency (ratio of actual and nominal concentration) was 23%.
No. of animals per sex per dose:
5 (five)
Control animals:
no
Sex:
female
Dose descriptor:
discriminating conc.
Effect level:
ca. 5.1 mg/L air (analytical)
Based on:
test mat.
95% CL:
>= 4.95 - <= 5.25
Exp. duration:
4 h

The time-weighted mean actual concentration was 5.1 ± 0.15 mg/L. The nominal concentration (amount of test item used divided by the volume of pressurized air used) was 22 mg/L. The generation efficiency (ratio of actual and nominal concentration) was 23%. The concentration was measured at time points (n=18) that were equally distributed over the exposure period, the results of which demonstrated that the item was sufficiently stable. The generation was interrupted on three occasions in order to remove test item deposits from the system. To compensate for these interruptions, the generation time was elongated by 4 minutes in order to achieve an actual exposure time of 240 minutes.

The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were

determined twice during the exposure period. The MMAD was 1.8 µm (gsd 2.1) and 1.4 µm (gsd 1.9).

Mortality

No mortality occurred.

Clinical Signs

No clinical signs indicating test item related toxicity were seen during and after exposure. Black and

blue staining by the test item of several body parts was seen throughout the observation period.

Body Weights

Overall body weight gain in males and females was within the range expected for rats of this strain

and age used in this type of study and were therefore considered not indicative of toxicity.

Macroscopic Findings

No abnormalities were found at macroscopic post mortem examination of the animals.

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The inhalatory LC50, 4h value of K1600 black dye in Wistar rats was established to exceed 5 mg/L

Based on these results K1600 black dye does not have to be classified and has no obligatory labelling
requirement for acute inhalation toxicity according to the Globally Harmonized System of Classification
and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments) and
Regulation (EC) No 1272/2008 on classification, labelling and packaging of items and mixtures
(including all amendments).
Executive summary:

The study was carried out based on the guidelines described in: - OECD Guidelines, Section 4, Health Effects. No.403, "Acute Inhalation Toxicity", Sep 2009. - Commission Regulation (EC) No 440/2008, B.2. Acute Toxicity (inhalation), L142, May 2008. - EPA OPPTS 870.1300, Acute inhalation Toxicity. EPA 712-C-98-193, August 1998. - JMAFF Guidelines (2000), including the most recent revisions. K1600 black dye was administered as an aerosol by nose-only inhalation for 4 hours to one group of five male and five female Wistar rats. Generation as a dust was technically not possible and therefore the test item was mixed with water and generated as a liquid aerosol. Mortality and clinical signs were observed daily during the observation period and body weights were determined on Days 1, 2, 4, 8 and 15. Macroscopic examination was performed after terminal sacrifice (Day 15). The time-weighted mean actual concentration was 5.1 ± 0.15 mg/L. The nominal concentration (amount of test item used divided by the volume of pressurized air used) was 22 mg/L. The generation efficiency (ratio of actual and nominal concentration) was 23%. The concentration was measured at time points (n=18) that were equally distributed over the exposure period, the results of which demonstrated that the item was sufficiently stable. The generation was interrupted on three occasions in order to remove test item deposits from the system. To compensate for these interruptions, the generation time was elongated by 4 minutes in order to achieve an actual exposure time of 240 minutes. The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were determined twice during the exposure period. The MMAD was 1.8 µm (gsd 2.1) and 1.4 µm (gsd 1.9). No mortality occurred. No clinical signs indicating test item related toxicity were seen during and after exposure. Black and blue staining by the test item of several body parts was seen throughout the observation period. Overall body weight gain in males and females was within the range as expected for rats of this strain and age used in this type of study. No abnormalities were found at macroscopic post mortem examination of the animals. The inhalatory LC 50, 4h value of K1600 black dye in Wistar rats was established to exceed 5 mg/L. Based on these results K1600 black dye does not have to be classified and has no obligatory labelling requirement for acute inhalation toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of items and mixtures (including all amendments).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating conc.
Value:
5 100 mg/m³

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:
08 April 2015 - 29 September 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This valid study was performed at a GLP laboratory in accorance with OECD, EU, USA and Japanese test guielines for the determination of dermal toxicity.
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
yes
Remarks:
Deviations from the maximum level of temperature daily mean relative humidity occurred. Laboratory historical data do not indicate an effect of the deviations. The study integrity was not adversely affected by the deviation.
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
yes
Remarks:
Deviations from the maximum level of temperature daily mean relative humidity occurred. Laboratory historical data do not indicate an effect of the deviations. The study integrity was not adversely affected by the deviation.
Qualifier:
according to
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Deviations:
yes
Remarks:
Deviations from the maximum level of temperature daily mean relative humidity occurred. Laboratory historical data do not indicate an effect of the deviations. The study integrity was not adversely affected by the deviation.
Qualifier:
according to
Guideline:
other: JMAFF Guidelines (2000), including the most recent revisions.
Deviations:
yes
Remarks:
Deviations from the maximum level of temperature daily mean relative humidity occurred. Laboratory historical data do not indicate an effect of the deviations. The study integrity was not adversely affected by the deviation.
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approx. 10 weeks old
- Weight at study initiation: Body weight variation did not exceed +/- 20% of the sex mean.
- Housing: Individually housed in labeled Makrolon cages ontaining sterilized sawdust as bedding material.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days before start of treatment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24°C
- Humidity (%): 40 to 70%
- Air changes (per hr): 10 hours
- Photoperiod (hrs dark / hrs light): 12-hour light/12-hour dark cycle
Type of coverage:
occlusive
Vehicle:
water
Details on dermal exposure:
TEST SITE
- Area of exposure: No data
- % coverage: 10% of the total body surface, i.e. approx. 25 cm² for males and 18 cm² for females.
- Type of wrap if used: consisting of a surgical gauze patch (Surgy 1D)*, successively covered with aluminum foil and Coban elastic bandage. A piece of Micropore tape* was additionally used for fixation of the bandages in females only.

REMOVAL OF TEST SUBSTANCE
- Washing: After which dressings were removed and the skin cleaned of residual test substance using tap water.
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2000 mg/kg body weight.
- Concentration (if solution) :10 mL/kg body weight.

VEHICLE
- Concentration (if solution): 20 to 200 mg/mL of test substance in vehicle
Duration of exposure:
24 hours
Doses:
2000 mg/kg body weight
No. of animals per sex per dose:
5 males and 5 females, one dose.
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Mortality and viability were observed twice daily. Body weights were examined on days 1, 8 and 15.
- Necropsy of survivors performed: yes, at the end of the study.
- Other examinations performed: clinical signs, body weight,organ weights and descriptions of all internal macroscopic abnormalities were
recorded.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality occurred.
Clinical signs:
Piloerection, ptosis and/or chromodacryorrhoea were noted for one male animal and three female animals on Day 1. Blue staining by the test substance was seen on the treated skin and tail of all animals throughout the observation period. Focal erythema or scales were seen in the treated skin area of three animals during the observation period.
Body weight:
The changes noted in body weight gain in males and females were within the range expected for rats used in this type of study and were therefore considered not indicative of toxicity.
Gross pathology:
No abnormalities were found at macroscopic post mortem examination of the animals.

 
Table 1: Mortality Data
Test Day 1 1 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Hours after treatment 0 2 4
Males 2000 mg/kg - - - - - - - - - - - - - - - - -
Females 2000 mg/kg - - - - - - - - - - - - - - - - -

 
Table 2: Clinical Signs
Test Day Max Grade 1 1 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Hours after treatment 0 2 4

Males 2000 mg/kg

Animal 1
Various:
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 2
Various:
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 3
Various:
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 4
Skin / fur
Erythema focal (treated skin) (4) - - - - - 1 1 1 - - - - - - - - -
Various:
Blue (tail) (1) - - - - 1 1 1 1 1 - - - - - - - -
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 5
Skin / Fur:
Piloerection (1) - - 1 - - - - - - - - - - - - - -
Scales (treated skin) (3) - - - - - 1 1 1 - - - - - - - - -
Various:
Ptosis (3) - - 1 - - - - - - - - - - - - - -
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Females 2000 mg/kg
Animal 6
Skin / Fur:
Piloerection (1) - - 1 - - - - - - - - - - - - - -
Secretion/ excretion
Chromodacryorrhoea (Snout) (3) - - 1 - - - - - - - - - - - - - -
Various:
Ptosis (3) - - 1 - - - - - - - - - - - - - -
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 7
Various:
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 8
Skin / Fur:
Piloerection (1) - - 1 - - - - - - - - - - - - - -
Various:
Ptosis (3) - - 1 - - - - - - - - - - - - - -
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 9
Skin / Fur:
Piloerection (1) - - 1 - - - - - - - - - - - - - -
Various:
Ptosis (3) - - 1 - - - - - - - - - - - - - -
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1
Animal 10
Skin / Fur:
Scales (treated skin) (3) - - - - - - - - - - - - - 1 1 - -
Various:
Blue (tail) (1) - - - - 1 1 1 1 1 1 1 1 1 1 1 1 1
Blue (Treated skin) (1) - - - 1 1 1 1 1 1 1 1 1 1 1 1 1 1

 
Table 3: Body Weights (gram)
Sex/ Dose level Animal Day 1 Day 8 Day 15
Males 2000 mg/kg
1 274 298 331
2 274 298 329
3 282 303 332
4 272 301 329
5 260 265 278
Mean 272 293 320
ST. DEV. 8 16 23
N 5 5 5
Females 2000 mg/kg
6 210 203 211
7 194 196 200
8 207 209 211
9 208 207 225
10 202 203 215
Mean 204 204 212
ST. DEV. 6 5 9
N 5 5 5
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The dermal LD50 value of K1600 black dye in Wistar rats was established to exceed 2000 mg/kg body weight. Based on these results, K1600 black dye does not have to be classified and has no obligatory labelling requirement for acute dermal toxicity according to the:
- Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments),
- Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).
Executive summary:

K1600 black dye was administered to five Wistar rats of each sex by a single dermal application at 2000 mg/kg body weight for 24 hours. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice (Day 15).

No mortality occurred. Piloerection, ptosis and/or chromodacryorrhoea were noted for one male animal and three female animals on Day 1.

The changes noted in body weight gain in males and females were within the range expected for rats used in this type of study and were therefore considered not indicative of toxicity.

No abnormalities were found at macroscopic post mortem examination of the animals.

The dermal LD50 value of K1600 black dye in Wistar rats was established to exceed 2000 mg/kg body weight.

Based on these results, K1600 black dye does not have to be classified and has no obligatory

labelling requirement for acute dermal toxicity according to the:

- Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments),

- Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating dose
Value:
2 000 mg/kg bw

Additional information

No adverse effects were observed in any acute toxicity study at the limit dose tested (2000 mg/kg oral, 2000mg/kg dermal or 5 mg/L inhalation)

Justification for classification or non-classification

No adverse effects were observed in any of the acute toxicity studies conducted at the limit dose. The criteria for classifcation were not met.