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Description of key information

LD50 oral > 2000 mg/kg bw

Dermal and inhalation exposure is unlikely, thus no acute toxicity value is available and no further investigation is required.

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 May 22th to June 6th, 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
2001
Qualifier:
according to guideline
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
2002
Qualifier:
according to guideline
Guideline:
other: Japan MAFF Testing Guideline of 12 Nosan no. 8147, 2000
GLP compliance:
yes (incl. QA statement)
Test type:
acute toxic class method
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Specie: Rattus norvegicus.
- Source: TECAM Animal Facility (S. Roque, SP).
- Age at study initiation: young adult rats 8 - 10 weeks old.
- Variation in weight: not greater than 20 % of the mean weight.
- Housing: 3 animals per cage.
- Diet: pelleted commercial diet (Biobase Biotec), ad libitum. Feed is analysed by testing laboratory periodically for microbiological contaminants
- Water: filtered water, ad libitum.
- Fasting period before study: animals were fasted approxirnately 14 hours prior to the test substance administration. Animals returned to ad Iibitum feeding apprornmately 3 hours after dosing.
- Acclimatization period: 5 days prior to dosing in a controlled room.
- Selection: animals exhibiting abnormal signs during the acclimatization period were not used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 - 23 °C
- Humidity: 62 %
- Photoperiod: 12 hours cycle dark/light
Route of administration:
oral: gavage
Vehicle:
water
Remarks:
deionized
Details on oral exposure:
Rationale for the selection of the starting dose: as test substance was suspected not to be toxic, 2000 mgkg bw was the selected dose for starting. The time interval between treatment groups was determined by the onset, duration and severity of toxic signs.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
Two groups of 3 females
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days.
- Frequency of observations and weighing: body weights were recorded shortly before administration, weekly thereafter and at the end of the study. Animals were observed individually after dosing during the first 24 hours with special attention given during the first 4 hours (day 1).
- Necropsy of survivors performed: at the end of 14 days, carbon dioxide was used for euthanasia of the animals. Gross pathology examination was performed for 100 % of the animals. Necropsy findings were registered for the intestinal tract (duodenum, jejunum, ileum, cecum) and the major organs such as liver, kidney, heart, spleen, Iymph nodes, respiratory tract (lungs, trachea, bronchi, diaphragm), thyroid, oesophgus, stomach, pancreas, muscles, bladder, uteus and gonads. After examination, animals were wrapped and disposed as a special residue of the Municipal Health Service.
- Other examinations performed: sign and symptoms were recorded at least once each workday for individual animals. A check for dead or moribund animals were made once at least once each workday and once on Saturdays, Sundays and on public holidays. Clinical observations included possible changes in skin and fur, eyes and mucous membranes, dyspnoea, changes in the behaviour, tremors, convulsions, salivation, diarrhea, prostration, coma and death.
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality was observed among the tested animals.
Clinical signs:
No treatment related signs were observed after a single administration.
Body weight:
Body weight changes were within the range of physiological variability.
Gross pathology:
No macroscopic changes were observed in any of the major organs of the examined animals.

Individual and group rnean body weight (bw) of animals treated at 2000 mg/kg bw of the test substance

Animals Bw (g)
Day 1 Day 7 Day 14 Bw changes
1 172.9 202.5 219.7 + 46.8
2 185.8 206.9 221.4 + 35.6
3 163.6 185.6 198.6. + 35.0
4 176.7 194.1 204.8 + 28.1
5 184.2 212.0 223.9 + 39.7
6 184.9 189.5 197.9 + 13.0
Mean 178.0 198.4 211.0 + 33.0

 

Individual clinical observation of animals treated at 2000 mg/kg bw of the test substance

Animals Days of observation
1 2 3 – 5 6 7 8 9 – 14
1 NA NA NA NA NA NA NA
2 NA NA NA NA NA NA NA
3 NA NA NA NA NA NA NA
4 NA NA NA NA NA NA NA
5 NA NA NA NA NA NA NA
6 NA NA NA NA NA NA NA

NA: no alterations

Individual necropsies of animals treated at 2000 mg/kg bw of the test substance

Macroscopic observations Females
1 2 3 4 5 6
Oesophagus NA NA NA NA NA NA
Trachea NA NA NA NA NA NA
Thyroid NA NA NA NA NA NA
Lungs NA NA NA NA NA NA
Bronchi NA NA NA NA NA NA
Heart NA NA NA NA NA NA
Diaphragm NA NA NA NA NA NA
Stomach NA NA NA NA NA NA
Intestine NA NA NA NA NA NA
Spleen NA NA NA NA NA NA
Pancreas NA NA NA NA NA NA
Liver NA NA NA NA NA NA
Lymph nodes NA NA NA NA NA NA
Muscles NA NA NA NA NA NA
Kidney NA NA NA NA NA NA
Bladder NA NA NA NA NA NA
Ovarium NA NA NA NA NA NA
Uterus NA NA NA NA NA NA

NA: no alterations

Interpretation of results:
other: not classified, according to the CLP Regulation (EC 1272/2008)
Conclusions:
LD50 > 2000 mg/kg bw
Executive summary:

The study was carried out to assess acute toxicity following a single oral administration to rats of the test substance according to the OECD guideline 423. Wistar rats (Rattus norvegicus) were selected and maintained under controlled environmental conditions. Based on individual body weight, dosing of the test substance was calculated in mg/kg body weight (bw). Two groups of three female rats received 2000 mg/kg bw. Animals were observed for 14 days.

At the end of this period, no mortality or signs of evident toxicity were observed. Gross pathology examination was performed on all tested animals. No macroscopic alterations were observed in any of the major organs of the examined animals. Under the test conditions, the LD50 was found to be greater than 2000 mg/kg bw. Therefore, the test substance is not acutely toxic via the oral route.

Conclusion

LD50 > 2000 mg/kg bw

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw

Acute toxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Justification for Read Across is explained in the endpoint summary and it is further detailed in the report attached to the IUCLID section 13.
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
GLP compliance:
yes (incl. QA statement)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Area of exposure: 6.0 cm × 5.0 cm (30.0 cm^2)
- Type of wrap if used: the test substance was transferred to a wet-gauze layer of a "Cutiplast steril®" and coated with air-tight "Leukoflex®". The gauze strip was placed on the rats back and secured in place using "Peha®-Haft" cohesive stretch tape and additionally covered with a "Lomir biomedical Inc rat jacket," connected with a safety pin to the stretch tape to ensure that animals could not ingest the test substance.

REMOVAL OF TEST SUBSTANCE
- Washing: dressings were removed and the area was rinsed with tepid water using soap and gently patting the area dry.
- Time after start of exposure: approximately 24 h.
Duration of exposure:
24 hours
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5 males and 5 females
Control animals:
not required
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No deaths occurred; the dose was tolerated by males and females.
Clinical signs:
No toxicologically relevant clinical signs.
Body weight:
No toxicological effects on weight development.
Gross pathology:
No findings.
Interpretation of results:
other: not classified, according to the CLP Regulation (EC 1272/2008)
Conclusions:
LD50 > 2000 mg/kg bw
Executive summary:

The acute dermal toxic potential of the test substance was examined according to the OECD guideline 402. Groups of 5 male and 5 female Wistar rats received a single dermal dose of 2000 mg/kg body weight of the test item applied semiocclusively for 24 hours. A dose of 2000 mg/kg body weight was tolerated by male and female rats without mortalities, toxicologically relevant clinical signs, toxicological effects on weight development and gross pathological findings.

Conclusion

LD50 > 2000 mg/kg bw

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw

Additional information

ACUTE TOXICITY - ORAL ROUTE

The oral acute toxicity of Basic Blue 140 (BBl140) was tested with a single administration to rats, according to the OECD guideline 423. Wistar rats (Rattus norvegicus) were selected and maintained under controlled environmental conditions. Two groups of three female rats received 2000 mg/kg bw. Animals were observed for 14 days. At the end of this period, no mortality or signs of evident toxicity were observed. Gross pathology examination was performed on all tested animals. No macroscopic alterations were observed in any of the major organs of the examined animals. The LD50 was found to be greater than 2000 mg/kg bw. Therefore, the test substance is not acutely toxic via the oral route.

ACUTE TOXICITY - INHALATION ROUTE

No acute toxicity studies by inhalation route are available on BBl140.

Nevertheless, because of the physical state and the trade forms of the substance under registration, inhalation is not an appropriate route of exposure. Acute toxicity results for the other exposure routes indicate no concern.

ACUTE TOXICITY - DERMAL ROUTE

The inhalation and the skin contact of BBl140 are unlikely. Furthermore, because of the physical/chemical properties of the substance, the dermal absorption is expected as negligible.

According to the Commission Regulation (EU) 2016/863, amending Annexes VII and VIII to REACH Regulation (EC 1907/2006) as regards skin corrosion/irritation, serious eye damage/eye irritation and acute toxicity, testing by the dermal route does not need to be conducted if no systemic effects have been observed in in vivo studies with dermal exposure (e.g. skin irritation, skin sensitisation). Furthermore, it is explained that recent scientific analysis of available data from in vivo acute toxicity studies have shown that substances that are not toxic via the oral route may be expected with high certainty to be also non-toxic via the dermal route. Therefore, testing those substances via the dermal route does not provide essential information for their safety assessment.

The amendment is the consequence of studies and scientific debates. In particular, the 15th Meeting of Competent Authorities for REACH and CLP (CARACAL, 2014) concluded that an adaptation of point 8.5.3 of Annex VIII to REACH is justified in order to not require information on acute dermal toxicity for substances that have shown no toxicity in acute oral toxicity test up to the limit dose of 2000 mg/kg bw.

Furthermore, the acute dermal toxic potential was investigated for the Similar Substance 01, which was examined according to the OECD guideline 402. Groups of 5 male and 5 female Wistar rats received a single dermal dose of 2000 mg/kg body weight of the test item applied semiocclusively for 24 hours. The dose administered was tolerated by male and female rats without mortalities, toxicologically relevant clinical signs, toxicological effects on weight development and gross pathological findings.

BBl140 and Similar Substance 01 share the same phthalocyaninic core and, in both cases, they present a zwitterionic moiety formed by sulphonic acid, which conducts an acid function, and a tertiary amine on the sulphonaminic chain, characterized by a basic function. BBl140 presents a further sulphonaminic functionalization, of which chain ends with a tertiary amine that, based on the stechiometrical ratio reported in the analytical characterization, in most of the cases can be salified with acetic acid. It is expected that this difference has not a significant impact on the acute toxicity potential by dermal route.

The extend of percutaneous absorption of a substance depends largely on the physical and chemical properties of the substance itself. In particular, factors like the degree of ionization, molecular size and water and lipid solubilities influence penetration through the skin.

Local factors such as the pH, ambient temperature, blood perfusion and local anatomy also play a role. Taking into account the molecular size and the hydrophilic behaviour of both BBl140 and Similar Substance 01, it is not expected that dermal absorption occurred.

The read across approach has been further detailed in the report attached to the IUCLID section 13.

REFERENCE

Bulgheroni A., Kinsner-Ovaskainen A., Hoffmann S., Hartung T., Prieto P. (2009). Estimation of acute oral toxicity using the No Observed Adverse Effect Level (NOAEL) from the 28 day repeated dose toxicity studies in rats. Regulatory Toxicology and Pharmacology 53, 16–19.

CARACAL (2014). 15th Meeting of Competent Authorities for REACH and CLP (CARACAL), 8 – 9 July 2014. Charlemagne building, Brussels, Belgium. Brussels, 26 July 2014. Doc. CA/61/2014. Stakeholder proposal to modify REACH standard information requirements for acute toxicity (REACH Annex VIII, point 8.5).

ECHA (2016). Guidance on Information Requirements and Chemical Safety Assessment Chapter R.7a: Endpoint specific guidance Draft Version 5.0 August 2016. Available: https://echa.europa.eu/documents/10162/13643/ir_csa_r7a_r7-4_caracal_en.pdf/1f97ae8b-052c-4713-959a-5fd742b63b03

Justification for classification or non-classification

According to the CLP Regulation (EC 1272/2008), 3.1 Acute toxicity section, substances can be allocated to one of four toxicity categories based on acute toxicity by the oral, dermal or inhalation route according to the numeric criteria. Acute toxicity values are expressed as (approximate) LD50 (oral, dermal) or LC50 (inhalation) values or as acute toxicity estimates (ATE).

The oral LD50 value was established to be greater than 2000 mg/kg body weight, therefore the test substance is out of any classification limit for acute oral toxicity (oral acute toxicity category 4: 300 < ATE ≤ 2000 mg/kg bw).

Dermal and inhalation exposure is unlikely, thus no acute toxicity value is available and no further investigation is required.

In conclusion, the test substance is non classified for oral acute toxicity, according to the CLP Regulation (EC 1272/2008).