Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Administrative data

Description of key information

Skin sensitisation (OECD 429): not sensitising

RA from source substance Bismuth hydroxide nitrate oxide (CAS 1304-85-4)

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
19 Mar - 23 April 2010
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:
22 July 2010
Deviations:
yes
Remarks:
different evaluation of cellular proliferation (cell count measurement instead of incorporation of 3H-methyl thymidine)
Qualifier:
according to guideline
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
6 Juli 2012
Deviations:
yes
Remarks:
different evaluation of cellular proliferation (cell count measurement instead of incorporation of 3H-methyl thymidine)
Qualifier:
according to guideline
Guideline:
other: Note for Guidance on Non-Clinical Tolerance Testing of Medicinal Products, CPMP/SWP/2145/00, adopted March 1, 2001.
Deviations:
not specified
GLP compliance:
yes (incl. QA statement)
Remarks:
Behörde für Soziales, Familie, Gesundheit und Verbraucherschutz, Freie und Hansestadt Hamburg
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
NMRI
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: NMRI mice supplied by Charles River Deutschland GmbH were used in this experiment.
- Age at study initiation: 8 weeks
- Weight at study initiation: 22 - 28 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 was used as bedding material for the cages. The cages were changed and cleaned once a week.
- Diet: ad libitum; commercial diet ssniff R/M-H V1534. Food residue was removed.
- Water: ad libitum
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 3°C (maximum range)
- Humidity: 55 ± 15% (maximum range)
- Photoperiod: 12/12 hours dark/light cycle
Vehicle:
other: a mixture of acetone / olive oil (3+1, v/v)
Concentration:
10%, 25% and 50% w/w
No. of animals per dose:
30 (5 groups of 6 female animals each)
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: The vehicle (a mixture of acetone/olive oil) was selected on the basis of maximising the test concentrations and solubility whilst producing a solution/suspension suitable for application of the test item.
- Irritation: 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 Bismuth hydroxide nitrate oxide in acetone/olive oil (3+1 v/v), w/w were examined.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
At the start of the experiment the mice were weighed and assigned to each of the 5 groups by a randomisation program (block size n = 6).
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: 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. An index above 1.4 is considered positive.
- Other: Furthermore, 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.

TREATMENT PREPARATION AND ADMINISTRATION:
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.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
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 would have been 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.
Positive control results:
The positive control group caused the expected increases in lymph node cell count (statistically significant at p ≤ 0.01). Even though also the lymph node weight was increased in the positive control (indicating factor to be a substance with irritating properties) the no observed “acute skin reaction” indicates that the positive control group has sensitising properties. The values for the stimulation index of lymph node cell count was 1.94. Therefore, the study could be regarded as valid.
Key result
Parameter:
SI
Value:
1.181
Test group / Remarks:
Bismuth hydroxide nitrate oxide at concentrations of 10%
Key result
Parameter:
SI
Value:
1.145
Test group / Remarks:
Bismuth hydroxide nitrate oxide at concentrations of 25%
Key result
Parameter:
SI
Value:
1.246
Test group / Remarks:
Bismuth hydroxide nitrate oxide at concentration of 50%
Remarks on result:
other:
Remarks:
Treatment with a concentration of 50% revealed indeed a statistical significantly increased value (SI of 1.246), but all stimulation indices for the lymph node cell count were beneath the threshold value of 1.4. Hence, the test item is classified as not sensitising. The threshold level for the ear weight of 1.1 was not exceeded and the lymph node weights were not increased in a dose-related way, i.e. no irritating properties were noted. A table with more details is attached to this endpoint study record.
Cellular proliferation data / Observations:
Treatment with the test item at concentrations of 10% or 25% did not reveal statistical significantly increased values for lymph node cell count. Treatment with a concentration of 50% revealed indeed a statistical significantly increased value, but all stimulation indices for the lymph node cell count were beneath the threshold value of 1.4. Hence, the test item is classified as not sensitising.
No signs of local or systemic intolerance were recorded. The animal body weight was not affected by the treatment.

See tables attached under 'background material'.

Summary

The purpose of this study was to determine the sensitising potential of Bismuth hydroxide nitrate oxide in the local lymph node assay in mice. The study was performed according to OECD 429.

Stimulation indices were calculated for the lymph node cell count, lymph node weight, ear weight and ear thickness by dividing the average values per group of the test item treated animals by the vehicle treated ones. Values above 1.4 (lymph node cell count to identify sensitisation) or 1.1 (ear weight to identify irritation) are considered positive (these values were fixed empirically during the inter-laboratory validation of this method. 3 concentrations of Bismuth hydroxide nitrate oxide (10%, 25% and 50%, w/w) suspended in acetone/olive oil (3+1, v/v) were tested in six female NMRI mice per group and compared to a vehicle control group. In addition, a positive control group (30% solution (v/v) of α-hexyl cinnamic aldehyde in acetone/olive oil (3+1, v/v)) was employed.

Treatment with Bismuth hydroxide nitrate oxide at concentrations of 10% or 25% did not reveal statistical significantly increased values for lymph node cell count. Treatment with a concentration of 50% revealed indeed a statistical significantly increased value, but all stimulation indices for the lymph node cell count were beneath the threshold value of 1.4. Hence, the test item is classified as not sensitising.

Interpretation of results:
other: CLP/ EU GHS criteria not met, no classification required according to Regulation (EC) No 1272/2008
Conclusions:
CLP: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Skin Sensitisation

There are in vivo data available regarding skin sensitisation for Bismuth chloride oxide (CAS 7787-59-9) in order to fulfil the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.3.

In addition read-across from an appropriate substance is conducted in accordance with Regulation (EC) No 1907/2006, Annex XI, 1.5. in order to fulfil the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.3. Structural similarities and similarities in properties and/or activities of the source and target substance are the basis of read-across. A detailed justification for the analogue read-across approach is provided in the technical dossier (see IUCLID Section 13).

For assessment of the skin sensitisation potential information will also be taken into account from the analogue substance Bismuth hydroxide nitrate oxide (CAS 1304-85-4) to fulfil the standard data requirements defined in Regulation (EC) No 1907/2006, Annex VIII, 8.3.

CAS 1304-85-4

A skin sensitisation test was performed with bismuth hydroxide nitrate oxide using the local lymph node assay in mice (reference 7.4.1 -1). The study was performed according to OECD 429 and GLP. In the main study, 6 female mice (per group) were treated with 25 μL of the appropriate dilution of the test item (10%, 25% and 50%, w/w), the vehicle alone or the positive control. This procedure was repeated the following two days and on Day 4, the animals were sacrificed. Other than inn OECD 429, stimulation indices were calculated for the lymph node cell count. Values above 1.4 (lymph node cell count to identify sensitisation) were considered positive. Treatment with the test item at concentrations of 10% and 25% did neither reveal statistical significantly increased values for lymph node cell count nor a SI value above the cut-off value of 1.4. Treatment with a concentration of 50% revealed a statistical significantly increased value, but it still below the cut-off value. Hence, the test item is classified as not sensitising.

This result is supported by 3 older studies conducted using the Landsteiner protocol and a protocol similar to the Landsteiner technique, respectively. These studies were performed with Bismuth chloride oxide (CAS 7787 -59-9). The testing protocol according to Landsteiner is an old technique that was never included in the original OECD TG 406. Since no data are available on positive control and challenge control substances, the reliability of these studies was set to 4, but they were sufficient in documentation to be used as supporting studies.

CAS 7787-59-9

A supporting study according to the modified Landsteiner technique (Landsteiner, K., and Jacob, J., Exptl. Med. 61, 643, 1935; Draize, J.R., Woodard, G., and Calvery, H.O., J. Pharmacol. Exptl. Therap. 82, 377, 1944) was performed with the test material (50% formulation) (reference 7.4.1 -2). In the main study, 8 guinea pigs were treated with the test material at 0.1% for intra- and epidermal induction, 3 times a week until a total of 9 injections. 4 animals served as the negative control group. 14 days after the induction, epidermal challenge was performed with 0.1% of the test material. 24 and 48 h after the challenge treatment skin examination revealed no skin reactions in the test group or in the control group. No information on positive control results is available.

Another supporting study according to the modified Landsteiner technique (Landsteiner, K., and Jacob, J., Exptl. Med. 61, 643, 1935; Draize, J.R., Woodard, G., and Calvery, H.O., J. Pharmacol. Exptl. Therap. 82, 377, 1944) was performed with the test material with albino guinea pigs (reference 7.4.1 -3). In the main study, 8 guinea pigs were treated with the test material at 0.1% for intra- and epidermal induction, 3 times a week until a total of 9 injections. 4 animals served as the negative control group. Epidermal challenge was performed with 0.05 mL of the test material (100%). Intradermal injections of the test material produced slight erythema within 24 h after most of the injections, which were attenuated after 48 h. The challenge dose produced only slight erythema within 24 h in 4/8 animals. The response area was decreased in 3/4 animals showing the positive response, in the other animal slightly increased. 2,4-dinitro-1-chlorobenzene (DNCB) (0.1% w/v) was used as positive control material in the study and several doses were tested in parallel to the test material. 24 and 48 h after the challenge treatment skin examination revealed slight erythema within 24 h after application of most doses of the positive control material, which remained until 48 h or were attenuated. Blanching was observed during week 2. The challenge dose produced slight erythema in all animals with an increase in the measured response of each. The response to the challenge dose was more pronounced after 24 h compared to the 48 h time point.

A skin sensitisation test was performed with the test substance (reference 7.4.1 -4). In the main study, 8 guinea pigs were injected with 0.05 mL of the test substance (0.1% in saline) for intradermal induction, followed by daily injections with 0.1 mL of the test substances until a total of 10 injections were reached. An intradermal challenge was performed 2 weeks after the last injection of the induction, using 0.5 mL of the test substance. Intradermal injections of the test substance produced small raised areas with a slight increase in colour compared to the surrounding area within 24 h. No positive control was tested.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Taken together, the available data on skin sensitisation with Bismuth hydroxide nitrate oxide (CAS1304-85-4) and on the target substance bismuth chloride oxide (CAS 7787-59-9) itself do not meet the criteria for classification according to Regulation (EC) No 1272/2008, and are therefore conclusive but not sufficient for classification.