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Administrative data

Description of key information

A GPMT was performed with the substance since the in vitro test methods and LLNA are not suitable for titanium containing substances. In the treated group (treatment dose of 50%), no macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase. In the control group (associated with the treatment dose of 50%), no macroscopic cutaneous intolerance reactions were recorded after the challenge phase.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
29 August 2016 to 31 October 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
yes
Remarks:
A relative humidity lower than 30% was registered from 28-31 Dec, from 3-9 Jan and on 19 Jan. The minimum value measured was 20%. This deviation is considered as without impact on the conclusion of the study.
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
In vitro and LLNA methods were determined not to be suitable for the titanium based test substance.
Specific details on test material used for the study:
• Sponsor’s identification: Dichlorobis(η-cyclopentadienyl)titanium (CAS# 1271-19-8)
• Batch No.: 0708501022
• Chemical name: bis(cyclopentadienyl) titanium dichloride
• CAS No.: 1271-19-8
• Date received: 16 Novembre 2016
• Storage: room temperature, darkness
• Container: smoked glass flask (n=1)
• Form: cristalline powder
• Quantity: 186.00 g (container + content)
• Colour: Red
• Production date: not specified
• Expiry date: 03 April 2019
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
Housing

The animals were housed in groups of 3 at the maximum in polycarbonate containers, the flooring of which was covered with dust-free cuttings and the top fitted with a stainless steel lid with a feeding device and drinking device of 500 mL.
The temperature and relative humidity of the main test were controlled to remain within target ranges of 19°C to 25°C and 30% to 70%, respectively.
The rate of air exchange was at least ten changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (07.00 to 19.00) and twelve hours darkness.

Food and drink
The drinking water (tap water from public distribution system) and food (ENVIGO, 2040C) were supplied ad libitum. Microbiological and chemical analyses of the water were carried out once every six months by Bureau Veritas – Eurofins (FRANCE).

Preparation of animals
Fifteen female albino guinea pigs of Dunkin-Hartley strain, supplied by Envigo (Kreuzelweg 53, 5961 NM HORST - The Netherlands). At the beginning of the main test, the animals were 4, 5 or 6 weeks old.

The animals were nulliparous and non-pregnant.

Prior to the test, the animals were kept for a minimum acclimatization period of 5 days, under stabling and nutritional conditions identical to those of the test.

Before the experimentation process, they were identified individually by marking with picric acid and by means of a numbered ring on the edge of one ear.

The animals were carefully shorn before each test item application:
- On the inter-scapular zone for the induction phase,
- On the dorso-lumbar zone for the challenge phase.

At least 3 hours before the first reading (challenge phase) they were shorn a second time in this dorso-lumbar zone.

The animals were weighed at the beginning of the test, after the second induction and at the end of the test.

Animal welfare
The standard study plan related to this study has been approved by the registered Ethics Committee No. 76.

The study was performed in accordance with the guidelines regarding the care and use of animals for experimental procedures:
- the European Communities Council Directive 2010/63/EU of 22 September 2010,
- the French Decree No. 2013-118 of 01 February 2013.

The animals were provided with suitable environmental enrichment (Tunnel).

The study was designed and was conducted to cause the minimum of suffering or distress to the animals, according to the guidelines and to our internal animal welfare’s procedure.
At the end of the study, the animals were euthanized by overdose of sodium pentobarbital.


Route:
intradermal
Vehicle:
olive oil
Concentration / amount:
GROUP 1 (control):
• 2 ID: Freund’s Complete Adjuvant diluted at 50 % in olive oil
• 2 ID: olive oil
• 2 ID: a mixture with equal volumes v/v :
- Freund’s Complete Adjuvant at 50% and olive oil

GROUP 2 (Treated):
• 2 ID: Freund’s Complete Adjuvant diluted at 50 % in olive oil
• 2 ID: test item at 2% in olive oil
• 2 ID: a test mixture in equal volumes v/v :
- Freund’s Complete Adjuvant at 50% and the test item at 4% in olive oil
Day(s)/duration:
7
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
other: Epricutaneous: Second Topical induction
Vehicle:
other: liquid paraffin
Concentration / amount:
GROUP 1 (control): 0.5 mL of liquid paraffin.
GROUP 2 (treated): 0.5 mL of the test item at 60% in liquid paraffin.
Day(s)/duration:
10
Adequacy of induction:
other: Day 10: Residue of orange coloration not preventing the erythema quotation was noted.
No.:
#20
Route:
epicutaneous, occlusive
Vehicle:
paraffin oil
Concentration / amount:
Day 21
The experimen0tal procedure of this phase was identical for both groups GROUP 1 (Control) and GROUP 2 (Treated) submitted to this experimentation: on the previously shorn dorso-lumbar zone, an application, under occlusive dressing, was performed during 24 hours:
- 1 sample cup containing the test item diluted at 50% (MNIC) and 1 sample cup containing the test item diluted at 25% in liquid paraffin (1/2 MNIC).
Day(s)/duration:
3
Adequacy of challenge:
other: Day 22 The occlusive dressing was removed and the treated areas were rinsed with liquid paraffin. Residue of orange coloration not preventing the erythema quotation was noted.
No. of animals per dose:
Main study

GROUP 1 (negative control): 5 female guinea pigs identified n° C8056 and n°C8066 to C8069

GROUP 2 (treated): 10 female guinea pigs identified n° C8070 to C8079

Details on study design:
Preliminary studies

Determination by intradermal injection of the Maximal Non Necrotizing Concentration (MNNC)

This test was conducted for the purpose of defining a MNNC of the test item which, on intradermal injection during the induction phase, does not risk causing too great a lesion (non-necrotizing concentration), should be well-tolerated systemically and should be the highest to cause mild-to-moderate skin irritation.


Two animals received a volume of 0.1 mL of the test item, on both sides of the spine, at the diluted concentration of 2% in olive oil in view to determine the MNNC.

A macroscopic evaluation of the cutaneous reactions was conducted 24 hours after the injections.


Determination by topical application of the Pre-Maximal Non Irritant Concentration
(Pre-MNIC)

This test, which allowed evaluating the irritancy potential of the test item, defined whether an application of sodium lauryl sulfate would be needed during topical induction phase.

The test item was applied on the dorso-lumbar zone of two guinea pigs shorn beforehand, with occlusive dressing for 24 hours, at 4 different concentrations: diluted at 60%, 30%, 20% and 10% in liquid paraffin.
After the removal of the occlusive dressing, the treated areas were rinsed with liquid paraffin.
Residue of yellow orange coloration not preventing the erythema quotation was noted.

A macroscopic evaluation of the cutaneous reactions was conducted 24 hours after removal of the dressing.


Determination by topical application of the Maximal Non Irritant Concentration (MNIC)

This test was carried out for the purpose of determining the MNIC of the test item without risk of an irritant effect during the challenge phase.

Three guinea pigs were treated according to the same treatment as animals from GROUP 1 (control) for the induction phase (i.e. olive oil and liquid paraffin).

During the challenge phase, the animals were treated with the test item placed onto the selected treatment sites and covered with an occlusive dressing for a period of 24 hours at 4 different concentrations: diluted at 30%, 20%, 10% and 5% in liquid paraffin.
After the removal of the occlusive dressing, the treated areas were rinsed with liquid paraffin.
Residue of yellow coloration not preventing the erythema quotation was noted.

In view of these results, the animals were treated at two additional concentrations: diluted at 50%, and 40% in liquid paraffin.
After the removal of the occlusive dressing, the treated areas were rinsed with liquid paraffin.
Residue of orange coloration not preventing the erythema quotation was noted.

A macroscopic evaluation of the cutaneous reactions was conducted 24 and 48 hours after removal of the occlusive dressing.

Challenge controls:

GROUP 1 (control): 0.5 mL of liquid paraffin.
Positive control substance(s):
yes
Remarks:
(Reference substance: alpha Hexylcinnamaldehyde Tests 30-32
Positive control results:
Please refer to the above section "Any other information on materials and methods"
Key result
Reading:
other: !st reading - Challenge phase
Hours after challenge:
24
Group:
test group
Dose level:
25% and 50% (Ten animals per dose group)
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase.
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
other: 2nd reading - challenge phase
Hours after challenge:
48
Group:
test group
Dose level:
25% and 50% (Ten animals per dose group)
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
In the treated group (treatment dose of 50%), no macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase.
Remarks on result:
no indication of skin sensitisation

ASSESSMENT OF THE SENSITISING POTENTIAL

Challenge phase

 

TEST ITEM:Dichlorobis(η-cyclopentadienyl)titanium (CAS# 1271-19-8)

 

Table 4

 

Application: topical under occlusive dressing

 

1stapplication date (D0):26 December 2016                      

- Preliminary studies

 

-MNNC determination:

24 hours after the injections, no cutaneous reaction was noted at the tested concentration of 2%.

The first induction of the Group 2 was carried out by intradermal injection at the maximal non necrosing concentration of 2% (Table 1).

 

-Pre MNIC determination:

24 hours after the removal of the occlusive dressings, discrete erythema was noted in one animal and no cutaneous reaction was noted in the other animal at the tested concentration of 60%.

No cutaneous reaction was noted in the other animals at the tested concentrations of 30%, 20% and 10% (Table 2).

 

In view of these results, the concentration selected was 60% for the 2ndinduction of the Group 2 and the MNIC determination began at the concentration of 30%.

 

-MNIC determination:

24 and 48 hours after the removal of the occlusive dressings, no cutaneous reaction was noted whatever the tested concentration (Table 3).

 

In view of this result, the concentrations selected were 50% (MNIC) and 25% (1/2 MNIC).

 

Main study

 

-Induction phase Group 2:

No cutaneous reaction was noted in all animals (10/10) 24 hours after the first induction.

Dryness of the skin was noted in all animals (10/10) 24 hours after the second induction. (Appendix 4)

 

-Induction phase Group 1:

No cutaneous reaction was noted in all animals (5/5) 24 hours after the first induction.

Dryness of the skin was noted in all animals (5/5) 24 hours after the second induction. (Appendix 4)

 

-Challenge phase Groups 1 & 2:

Overall results of the challenge phase with the test item (readings at 24 and 48 hours) are givenin Table 4.

Individual scores of macroscopic evaluations performed during challenge phase with the test item are given inTable 5.

 

In the treated group (treatment dose of 50%), no macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase.

In the control group (associated with the treatment dose of 50%), nomacroscopiccutaneous intolerance reactions were recorded after the challenge phase.

 

In the treated group (treatment dose of 25%), no macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase.

In the control group (associated with the treatment dose of 25%), nomacroscopiccutaneous intolerance reactions were recorded after the challenge phase.

 

 

Weight evolution

 No abnormality was recorded in the body weight gain of both groups.

 

 

Mortality

No mortality was registered during the main test.

 

 

Pathology

 

A necropsy was performed on the surviving animals.At the end of the test, the animals are anaesthetised with sodium pentobarbital and administration continued to fatal levels.

 

An external examination and opening of the abdominal and thoracic cavities for examinations of major organs were performed. The appearance of any macroscopic abnormalities was recorded, see Table 8.

 

A hard lump was noted under the right leg after the challenge phase in 50% (5/10) of the animals of treated group.

The lumps were collected on day 25 and were preserved in AFA (alcohol, formalin, and acetic acid).

Furthermore, the treatment site of the 1stinduction were collected and preserved in AFA for all animals (5 animals from negative control + 10 animals from treatment group).

Interpretation of results:
GHS criteria not met
Conclusions:
In view of these results, under these experimental conditions, the test item Dichlorobis(η-cyclopentadienyl)titanium (CAS# 1271-19-8) does not have to be classified in category 1 as a skin sensitizer, in accordance with the Regulation EC No. 1272/2008 on classification, labelling and packaging of substances and mixtures.
No signal word or hazard statement is required.
Executive summary:

The aim of the study was to evaluate the possible allergenic activity of the test item after intradermal and topical administration in guinea pigs.

 

According the results of the pretests, the induction phase (intradermic injection at 2% and topical application at 60%) was conducted with the test itemDichlorobis(η-cyclopentadienyl)titanium (CAS# 1271-19-8)to 10 Guinea pigsand a 10-day rest phase. The challenge phase conducted under occlusive dressing for 24 hours, consisted of a single topical application of the test item diluted at 50% and 25% in liquid paraffin. The experimental protocol was established according to the Magnusson and Kligman method (J. Invest. Dermatol. 1969. 52, 268-276) and in accordance with O.E.C.D. Test Guideline No.406 of July 17th, 1992, and the test method B.6 of council regulation No. 440/2008.

 

 

In the treated group (treatment dose of 50%), no macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase.

In the control group (associated with the treatment dose of 50%), nomacroscopiccutaneous intolerance reactions were recorded after the challenge phase.

 

In the treated group (treatment dose of 25%), no macroscopic cutaneous reactions attributable to allergy were noted after the challenge phase.

In the control group (associated with the treatment dose of 25%), nomacroscopiccutaneous intolerance reactions were recorded after the challenge phase.

 

A hard lump was noted under the right leg after the challenge phase in 50% (5/10) of animals in the treated group.

 

 

In conclusion, in view of these results, under these experimental conditions, the test itemDichlorobis(η-cyclopentadienyl)titanium (CAS# 1271 -19-8)does not have to be classified in category 1as a skin sensitizer,in accordance with the Regulation EC No. 1272/2008 on classification, labelling and packaging of substances and mixtures.

No signal word or hazard statement is required.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Based on the results from a GPMT the substance is not classified as a sensitizer.