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Description of key information

Skin sensitisation: not sensitising (modified OECD 429; method according to Ehlings et al. 2005; GLP compliant) (read across to cobalt zinc aluminate blue spinel)

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
According to the ECHA Guidance Read-Across Assessment Framework (RAAF, March 2017), the read-across hypothesis for systemic effects is based on the assumption that different compounds give rise to the same common moieties (“(Bio)transformation to common compound(s)”) to which the organisms are exposed.
A sufficient solubility in water is required to enable partition from the stratum corneum into the epidermis. Since the Pigments are nearly insoluble, dermal uptake is likely to be low. Additionally, neglectable leaching in artificial sweat was reported in the respective bioaccessability studies. Thus, dermal uptake of ingredients of the pigment (metals) is also expected to be low.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target substance Cobalt aluminate blue spinel is a complex inorganic colored pigment (CICP) and considered to be a UVCB under REACH, following the sector specific guidance for complex inorganic colored pigments (Eurocolour, 2017). Cobalt and aluminium expressed as the respective oxides are the main contributing elements. It is noted, that during the manufacture of CICPs chemical bonds of the raw materials are broken, and atoms are homogeneously and ionically interdiffused to form a specific crystalline matrix. Therefore, metals and oxygen atoms occupy specific places within the crystalline lattice, and individual oxides do not exist.
A detailed characterisation of the target substance is given in the justification document, attached in IUCLID section 13.2.

The source substance Cobalt zinc aluminate blue spinel is also a complex inorganic colored pigment (CICP) and considered to be a UVCB under REACH, following the sector specific guidance for complex inorganic colored pigments (Eurocolour, 2017). It consists of almost the same contributing elements (Cobalt, zinc and aluminium expressed, as oxides are the main constituent). It is noted, that during the manufacture of CICPs chemical bonds of the raw materials are broken, and atoms are homogeneously and ionically interdiffused to form a specific crystalline matrix. Therefore, metals and oxygen atoms occupy specific places within the crystalline lattice, and individual oxides do not exist.
A detailed characterisation of the source substance is given in the justification document, attached in IUCLID section 13.2.

The common characteristic of the target substance and the source substance is that both substances crystallise in the spinel structure, which forms a strong and inert crystal lattice. Both substances consist of almost the same chemical constituents. Furthermore, both show a similar, very low solubility in different artificial and aqueous media which is used for (eco)toxicological predictions.

3. ANALOGUE APPROACH JUSTIFICATION
Inorganic pigments like the target and source substance are characterised by inertness as a consequence of the specific synthesis process (calcination at high temperatures, approximately 1000°C), rendering the substance to be of a unique, stable crystalline structure in which all atoms are tightly bound within the lattice.

Upon dissolution the target as well as the source substances liberate cobalt and aluminium cations to a similar very low extent.

According to the OECD Guidance on grouping of chemicals (2014) the following basis for grouping of metal compounds can be assumed:
“The main assumption underlying the grouping of metal compounds is that toxicological and ecotoxicological properties are likely to be similar or follow a similar pattern as a result of the presence of a common metal ion (or ion complex including a hydrated metal ion). It is the bioavailability of the metal ion (or a redox form of this ion) at target sites that, besides the toxicity potency, will determine the occurrence and severity of the effects to be assessed. This is a reasonable assumption for the majority of inorganic compounds and some organic compounds (e.g. metal salts of some organic acids), in the absence of demonstrated relative differences in bioavailability.”

According to the ECHA Guidance Read-Across Assessment Framework (RAAF, March 2017), the read-across hypothesis for systemic effects is based on the assumption that different compounds give rise to the same common moieties (“(Bio)transformation to common compound(s)”) to which the organisms are exposed.

Since the target substance and the source substance release the same (eco-)toxicological relevant units under environmental/physiological relevant conditions, the overall ecotoxicity/systemic toxicity of the CICPs can be interpolated by assessing the (eco-)toxicity of the individual dissolved contributing elements. The analogue hypothesis, i.e. release of the common (eco-)toxicological units, applies to the target substance and the source substances.

However, acc. to ECHA Guidance Chapter R.6 – QSAR and Grouping of chemicals (2008) some metal containing UVCB compounds may not be appropriate for consideration in a category approach, as their effects will not be expected to be adequately described by their metal content. In fact read-across based solely on the metal content of the inorganic pigment Cobalt aluminate blue spinel would be highly overestimating, due to the very low bioavailability.

Transformation/Dissolution data ( see also Table 5 and 6 of the justification document)
The target substance showed a solubility in the transformation/dissolution test acc. to OECD 29 below 0.5 µg/L for the element Co after 24h, 7 days and 28days at pH6 and pH8 and a 1 mg/L loading. Aluminium was soluble up to 2 µg/L after 28 days at pH8 and up to 1.2 µg/L after 28 days at pH6.
The source substance showed a higher solubility in the transformation/dissolution test acc. to OECD 29 for the element aluminium and a comparable lower dissolution for the elements cobalt and zinc below the LOD after 7 and 28 days at pH6 and a 1 mg/L loading (Grané, 2010).

Bioaccessibility data (see also Table 7 and 8 of the justification document)
Bioaccessibility has been investigated experimentally in vitro by simulating dissolution under physiological conditions considered to mimic the most dermal exposure route: artificial sweat (ASW, pH 6.5) which simulates the hypoosmolar fluid, linked to hyponatraemia (loss of Na+ from blood), which is excreted from the body upon sweating.
The target as well as the source substance show a similarly low solubility of the individual elements in artificial sweat solution and aqueous media corroborating the inertness of the substances. The source substance shows slightly higher bioaccessibility after 24h for cobalt without showing any signs of systemic or local toxicity in a skin sensitisation study.

A slightly higher release of aluminium in artificial sweat solution is noted for the target substance. However, this is not relevant for the endpoint in question as soluble aluminium ions are not known for a skin sensitising potential:

“Aluminium is not regarded as a skin sensitiser. Aluminium chloride was tested in a murine local lymph node assay (LLNA) at doses up to 25% and there were no indications of a skin sensitisation potential (Basketter et al., 1999). A guinea pig maximisation test (GPMT) for aluminium chlorohydrate (ACH) dosed at 25%, found in the European Chemicals Agency database (ECHA, 1998), indicates that this substance is not sensitising. In addition, there is considerable history of use of aluminium containing cosmetic products with no indication in humans that aluminium is sensitising (AFSSAPS, 2011). In a few instances, sensitisation has been reported following application of aluminium compounds in children with a history of atopy (Goiset et al., 2018). The SCCS agrees that the available animal studies show that aluminium compounds used in antiperspirants are not skin sensitising. There is limited evidence that aluminium compounds can cause contact allergy in humans. However, taking into account the widespread use of these compounds, the SCCS considers this to be a rare phenomenon.” (SCCS, 2020)

Thus, based on the above-mentioned similarities the read across for the endpoint skin sensitization from Cobalt zinc aluminate blue spinel to Cobalt aluminate blue spinel is considered to be acceptable.

4. DATA MATRIX
A data matrix is presented in Appendix 1 of the read-across document (attached in IUCLID section 13.2).

A detailed read-across justification is provided in the read-across document (attached in IUCLID section 13.2) .

Reference:
SCCS (Scientific Committee on Consumer Safety) - Opinion on the safety of aluminium in cosmetic products - Submission II, 2020, SCCS/1613/19

Reason / purpose for cross-reference:
read-across source
Key result
Parameter:
SI
Remarks:
lymph node cell count
Value:
1.175
Test group / Remarks:
10 % w/w test item
Remarks on result:
other: SI: 1.136 (lymph node weight)
Key result
Parameter:
SI
Remarks:
ear weight
Value:
0.953
Test group / Remarks:
10 % w/w test item
Remarks on result:
other: SI: 1.056 (ear thickness)
Key result
Parameter:
SI
Remarks:
lymph node cell count
Value:
1.322
Test group / Remarks:
25 % w/w test item
Remarks on result:
other: SI: 1.273 (lymph node weight)
Key result
Parameter:
SI
Remarks:
ear weight
Value:
0.871
Test group / Remarks:
25 % w/w test item
Remarks on result:
other: SI: 1.103 (ear thickness)
Key result
Parameter:
SI
Remarks:
lymph node cell count
Value:
1.267
Test group / Remarks:
50 % w/w test item
Remarks on result:
other: SI: 1.386 (lymph node weight)
Key result
Parameter:
SI
Remarks:
ear weight
Value:
1.006
Test group / Remarks:
50 % w/w test item
Remarks on result:
other: SI: 1.125 (ear thickness)
Interpretation of results:
GHS criteria not met
Remarks:
Cobalt zinc aluminate blue spinel has been tested negative for skin sensitisation, and does not require classification as a skin sensitiser according to Directive 67/548/EEC and its subsequent amendments, and according to Regulation (EC) No 1272/2008 and subsequent regulations. Due to the analogy of the source and target substance it is considered justified to conclude that cobalt aluminate blue spinel likewise is void of skin sensitising properties, and does not require classification for skin sensitisation.
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-06-11 to 2010-07-15
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
2002-04-24
Deviations:
yes
Remarks:
, modified OECD 429 ,method according to Ehlings et al. 2005
Principles of method if other than guideline:
The test was performed in accordance with the method according to Ehling et al (2005): An European inter-laboratory validation of alternative endpoints of the murine local lymph node assay: first round, Toxicology 212 (2005) 60-68 and Ehling et al (2005): An European inter-laboratory validation of alternative endpoints of the murine local lymph node assay: 2nd round, Toxicology 212 (2005) 69-79.

Threshold values of the stimulation indices of lymph node cell count and ear weight were calculated by dividing the average values per group of the test item treated animals by the vehicle treated ones. Values above 1.4 (cell count) or 1.1 (ear weight) are considered positive
(these values were fixed empirically during the inter-laboratory validation of this method). In addition, the lymph node weights were determined for concentration related properties.
GLP compliance:
yes (incl. QA statement)
Remarks:
signed 2009-11-12
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
NMRI
Sex:
female
Details on test animals and environmental conditions:
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, 97633 Sulzfeld, Germany
- Age at study initiation: 9 weeks
- Weight at study initiation: 26 -31 g
- Housing: The animals were kept singly in MAKROLON cages (type III) with a basal surface of approx. 39 cm X 23 cm and a height of approx. 15 cm. Granulated textured wood (Granulat A2, J. Brandenburg, 49424 Goldenstedt, Germany) was used as bedding material for the cages.
- Diet (ad libitum): Commercial diet ssniff® R/M-H V1534 (ssniff Spezialdiäten GmbH, 59494 Soest, Germany)
- Water (ad libitum): Tap water
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22 +/- 3 °C (maximum range)
- Relative humidity: 55% +/- 15 % (maximum range)
- Air changes: 12 -18 times per hour
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
other: acetone/olive oil (3 + 1, v/v)
Concentration:
10 % w/w, 25 % w/w, and 50 % w/w of Pigment Al-Co-Zn
No. of animals per dose:
6 female mice
Details on study design:
RANGE FINDING TESTS:
A preliminary experiment was carried out in 3 animals to examine the irritating potential and handling/application of the test item in order to select the appropriate concentrations. Three concentrations of 10%, 25% and 50% of Pigment Al-Co-Zn in acetone/olive oil (3+1 v/v), w/w, were examined.
Results:
In a preliminary experiment, concentrations of 10%, 25% and 50%, employing 1 animal per concentration, were examined. No pronounced irritating properties were observed in this preliminary experiment at concentrations of 10%, 25% or 50%, no differences in ear weight and ear thickness were noted.

MAIN STUDY:
The test item was suspended in a mixture of acetone (Batch no. 32201, SIGMA ALDRICH Chemie GmbH, 82024 Taufkirchen, Germany) / olive oil (Batch no. 4602602, Henry Lamotte GmbH, 28197 Bremen, Germany) (3+1, v/v). The vehicle was selected on the basis of maximising the test concentrations and solubility whilst producing a solution/suspension suitable for application of the test item.

The test item suspension was administered to the dorsum of both animal's ears at an application volume of 25 µL/ear.

The experimental schedule of the assay was as follows:
Day 1:
The weight of each animal was individually identified and recorded. In addition, ear swelling measurements were carried out at the helical edge of both ears using an Oditest micrometer.
Open application of 25 µL of the appropriate dilution of the test item, the vehicle alone or the positive control (as appropriate) were administered to the dorsum of each ear.
Days 2 and 3:
The application procedure carried out on day 1 was repeated.
Day 4 (24 hours after the last application the animals were sacrificed under ether anaesthesia by cutting the aorta abdominalis):
Ear swelling measurements (immediately before sacrificing the mice) were carried out at the helical edge of both ears using an Oditest micrometer.
Punch biopsies of 8 mm in diameter of the apical area of both ears were prepared and immediately weighed on an analytical balance.
Lateral pairs of auricular lymph nodes draining the ear tissue were excised, carefully separated from remaining fatty tissue and weighed on an analytical balance immediately following preparation. The lymph nodes were then stored on ice in PBS /0.5% BSA and subjected to the preparation of single cell suspensions by mechanical tissue disaggregation. The cells were counted automatically in a cell counter.

OBSERVATIONS:
The following observations were made during the course of the study:

- Clinical signs: Animals were observed once daily for any clinical signs of local systemic irritation at the application site or of systemic toxicity. Observations were recorded for each individual animal. Cageside observations included skin/fur, eyes, mucous membranes, respiratory and circulatory systems, somatomotor activity and behaviour patterns. The onset, intensity and duration of any signs observed were recorded.
In addition, animals were checked regularly throughout the working day from 7:30 a.m. to 4:30 p.m. On Saturdays and Sundays animals were checked regularly from 8:00 a.m. to 12:00 noon with a final check performed at approximately 4:00 p.m., if applicable.
- Body weight: The weight of each mouse was recorded at the time of allocation of animals to groups (test day 1) and at the time of necropsy (test day 4).

ANALYSIS OF RESULTS:
The so-called stimulation (or LLN-) indices to determine the sensitising potential were calculated by dividing the average absolute lymph node weight or lymph node cell counts per group of the test item treated animals by the vehicle treated ones.
Thus, in case of no stimulating effect the index for the lymph node cell count is always below 1.4 (cut-off value). An index above 1.4 is considered positive.
For lymph node weight significance at p≤ 0.01 is considered positive (U-test according to MANN and WHITNEY). A possible concentration-response-relationship for the lymph node weight in order to determine a possible sensitising potential was examined by linear regression analysis employing PEARSON's correlation coefficient. Outliers were determined according to the Nalimov test.
In addition, the acute inflammatory skin reaction (irritating potential) was measured by ear weight determination of circular biopsies of the ears and ear thickness measurements on test day 1 and test day 4 to identify skin irritation properties of the test item employing the U-test according to MANN and WHITNEY by comparing the test groups to the vehicle control.
The stimulation indices were calculated by dividing the average ear weight and average ear thickness on test day 4 per group of the test item treated animals by the vehicle treated ones. The cut-off threshold value for ear weight was set at 1.1.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Please refer to "details on study design"
Positive control results:
The positive control group caused the expected increases in lymph node cell count and lymph node weight (statistically significant at p ≤ 0.01). The values for the stimulation index of lymph node cell count and lymph node weight were 1.81 and 1.77, respectively. Therefore, the study could be regarded as valid.
Ear weight: SI: 1.018
Key result
Parameter:
SI
Remarks:
lymph node cell count
Value:
1.172
Test group / Remarks:
10 % w/w test item
Remarks on result:
other: SI: 1.136 (lymph node weight)
Key result
Parameter:
SI
Remarks:
ear weight
Value:
0.953
Test group / Remarks:
10 % w/w test item
Remarks on result:
other: SI: 1.056 (ear thickness)
Key result
Parameter:
SI
Remarks:
lymph node cell count
Value:
1.322
Test group / Remarks:
25 % w/w test item
Remarks on result:
other: SI: 1.273 (lymph node weight)
Key result
Parameter:
SI
Remarks:
ear weight
Value:
0.871
Test group / Remarks:
25 % w/w test item
Remarks on result:
other: SI: 1.103 (ear thickness)
Key result
Parameter:
SI
Remarks:
lymph node cell count
Value:
1.267
Test group / Remarks:
50 % w/w test item
Remarks on result:
other: SI: 1.386 (lymph node weight)
Key result
Parameter:
SI
Remarks:
ear weight
Value:
1.006
Test group / Remarks:
50 % w/w test item
Remarks on result:
other: SI: 1.125 (ear thickness)
Cellular proliferation data / Observations:
DETAILS ON STIMULATION INDEX CALCULATION
Threshold values of the stimulation indices of lymph node cell count and ear weight were calculated by dividing the average values per group of the test item treated animals by the vehicle treated ones. Values above 1.4 (cell count) or 1.1 (ear weight) are considered positive (these values were fixed empirically during the inter-laboratory validation of this method).

RESULTS ON SKIN SENSITISATION
Treatment with Pigment Al-Co-Zn at concentrations of 10%, 25% or 50% did not reveal statistical significantly increased values for lymph node cell count. The stimulation indices of the lymph node cell count did not exceed the threshold level of 1.4. In addition, the lymph node weight was not increased significantly. Hence, the test item is classified as not sensitising.
The threshold level for the ear weight of 1.1 was not exceeded, i.e. no irritating properties were noted.
(Please refer to "Attached background material" for the raw data on stimulation indices)

CLINICAL OBSERVATIONS:
No signs of local or systemic intolerance were recorded.

BODY WEIGHTS:
The animal body weight was not affected by the treatment.
Interpretation of results:
GHS criteria not met
Conclusions:
Under the present test conditions, Pigment Al-Co-Zn at concentrations of 10%, 25% or 50% (w/w) in acetone/olive oil (3+1, v/v) did not reveal any sensitising properties in the local lymph node assay. Therefore, the test item must not be classified and labelled according to regulation (EC) No.: 1272/2008.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Read-across argumentation: due to the structural similarity of cobalt aluminate blue spinel and cobalt zinc aluminate blue spinel(both pigments are spinels with Co2+in octahedral lattice position; both pigments consist of the same components: Co, Al), as well as their similar physico-chemical properties (solubility) and their bioaccessibility, read-across is


 


Skin sensitisation


Read-across is performed from cobalt zinc aluminate blue spinel to cobalt aluminate blue spinel. Cobalt zinc aluminate blue spinel has been tested negative for skin sensitisation, and does not require classification as a skin sensitiser according to Directive 67/548/EEC and its subsequent amendments, and according to Regulation (EC) No 1272/2008 and subsequent regulations.


 


Hence, it is considered justified to conclude that cobalt aluminate blue spinel likewise is void of skin sensitising properties, and does not require classification for skin sensitisation.

Respiratory sensitisation

Endpoint conclusion
Additional information:

Respiratory sensitisation


There is no specific indication whatsoever to assume any potential for respiratory hypersensitivity of cobalt aluminate blue spinel for the following reasons:


 


(1) during long-lasting industrial practice in the inorganic pigment industry, no cases of hypersensitivity have been observed until now by workers exposed directly to the substance


(2) in contrast to bioavailable cobalt substance which are known to elicit sensitising properties, cobalt aluminate blue spinel is considered an inert, non-bioaccessible substance chemically and structurally very similar to cobalt zinc aluminate blue spinel which has been tested negative for skin sensitisation (LLNA assay)


(3) bioaccessibility data for both substances confirm that the substance cobalt aluminate blue spinel can be considered as inert (similar to cobalt zinc aluminate blue spinel), and shows very limited solubility in physiological media; in particular with respect to the release of cobalt under physiological conditions, the release is even lower than that documented for cobalt zinc aluminate blue spinel.


(4)  Assuming that both effects (skin and respiratory sensitisation) for cobalt are based on similar immunological mechanisms, the absence of skin sensitisation properties supports the assumption of an absence of respiratory sensitisation.


In conclusion, the classification criteria acc. to regulation (EC) 1272/2008 as respiratory sensitiser are not met for the pigment cobalt aluminate blue spinel.


No cases of hypersensitivity have been observed until now by workers exposed directly to the substance and bioaccessibility data for both substances (cobalt zinc aluminate blue spinel and cobalt aluminate blue spinel) confirm that the substance cobalt aluminate blue spinel can be considered as inert (similar to cobalt zinc aluminate blue spinel), and shows very limited solubility in physiological media; in particular with respect to the release of cobalt under physiological conditions, the release is even lower than that documented for cobalt zinc aluminate blue spinel. Assuming that both effects (skin and respiratory sensitisation) for cobalt are based on similar immunological mechanisms, the absence of skin sensitisation properties supports the assumption of an absence of respiratory sensitisation.


Hence,  cobalt aluminate blue spinel does not meet the criteria as respiratory sensitiser and should not be classified as such.

Justification for classification or non-classification

Skin sensitisation


Read across is made to cobalt zinc aluminate blue spinel. Cobalt zinc aluminate blue spinel has no skin sensitisation potential and does not require classification as skin sensitiser according to Regulation (EC) No 1272/2008 and subsequent regulations. Hence, cobalt aluminate blue spinel will also not be classified.


 


Respiratory sensitisation


During long-lasting industrial practice in the inorganic pigment industry, no cases of hypersensitivity have been observed until now by workers exposed directly to the substance. In contrast to bioavailable cobalt substance which are known to elicit sensitising properties, cobalt aluminate blue spinel is considered an inert, non-bioaccessible substance chemically and structurally very similar to cobalt zinc aluminate blue spinel which has been tested negative for skin sensitisation (LLNA assay). Bioaccessibility data for both substances confirm that the substance cobalt aluminate blue spinel can be considered as inert (similar to cobalt zinc aluminate blue spinel), and shows very limited solubility in physiological media; in particular with respect to the release of cobalt under physiological conditions, the release is even lower than that documented for cobalt zinc aluminate blue spinel.


 


Assuming that both effects (skin and respiratory sensitisation) for cobalt are based on similar immunological mechanisms, the absence of skin sensitisation properties supports the assumption of an absence of respiratory sensitisation.


 


In conclusion, the classification criteria acc. to regulation (EC) 1272/2008 as respiratory sensitiser are not met for the pigment cobalt aluminate blue spinel.