Hexaphenoxycyclotriphosphazene

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Version / remarks:

22 July 2010

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

482-200-1

EC Name:

-

Cas Number:

1184-10-7

Molecular formula:

C36H30N3O6P3

IUPAC Name:

hexaphenoxy-1,3,5,2λ⁵,4λ⁵,6λ⁵-triazatriphosphinine

Test material form:

solid

Specific details on test material used for the study:

Batch/Lot number: 190202
Description: White or light yellow crystal
Purity: 99.53%
Expiry date: 31 January 2021
Storage conditions: Controlled room temperature (15-25 ºC, below 70% relative humidity).
Safety precautions: Routine safety precautions (lab coat, gloves, safety glasses, face mask) for unknown materials were applied to assure personnel health and safety.

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

Species and strain: CBA/CaCrl mice
Source: Charles River UK Limited, Manston Road, Margate Kent, CT9 4LT, United Kingdom
Hygienic level: SPF at arrival; standard housing conditions during the study
Justification of strain: On the basis of OECD Guideline, mice of CBA/Ca or CBA/J strain can be used. Females are used because the existing database is predominantly based on females.
Number of animals: 4 animals / group
Sex: Female, nulliparous, non-pregnant
Age of animals at starting: 8 weeks old (age-matched, within one week)
Body weight range at starting: 18.0 – 19.8 grams (The weight variation in animals in the study did notexceed ± 20% of the mean weight.)
Acclimatisation time: 13 days
Animal health: Only healthy animals were used for the study. Health status was certified by the veterinarian.
Housing / Enrichment: Group caging / Additional enrichment (hiding tunnels) wasalso used during the study.
Cage type: Type II. polypropylene / polycarbonate
Bedding and nesting: Bedding and nesting materials were available to animalsduring the study
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 20.1 – 23.8°C
Relative humidity: 22 - 73%
Ventilation: 15-20 air exchanges/hour

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

10%, 5%, 2.5%

No. of animals per dose:

4

Details on study design:

In the main assay, twenty female CBA/CaCrl mice were allocated to five groups, each group comprised four animals:
- three groups of animals received HPCTP (formulated in DMF) at 10, 5, and 2.5% (w/v), respectively,
- a negative control group received the vehicle (DMF) only,
- a positive control group received 25% (w/v) HCA (dissolved in DMF).

The test item solutions were applied to the dorsal surface of the ears of the experimental animals (25 µL/ear) for three consecutive days (Days 1, 2 and 3) and then maintained on study for an additional 3 days. Cell proliferation in the (local) lymph nodes was assessed by measuring disintegrations per minutes after the incorporation of tritiated methyl thymidine (3HTdR) into the lymph nodes and the values obtained were used to calculate stimulation indices (SI) in comparison with the control group.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:

Positive controls responded as expected

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

ca. 0.6

Test group / Remarks:

HPCTP 10%(w/v) in DMF

Remarks on result:

no indication of skin sensitisation based on QSAR/QSPR prediction

Results 2

Key result

Parameter:

SI

Value:

ca. 1

Test group / Remarks:

HPCTP 5%(w/v) in DMF

Remarks on result:

no indication of skin sensitisation based on QSAR/QSPR prediction

Results 3

Key result

Parameter:

SI

Value:

ca. 1

Test group / Remarks:

HPCTP 2.5%(w/v) in DMF

Remarks on result:

no indication of skin sensitisation based on QSAR/QSPR prediction

Any other information on results incl. tables

No test item residue was observed in any of the animals. There were no indications of any irritancy at the site of application. No marked body weight losses (≥5%) were observed during the study in any of the animals. The results of the proliferation assay are summarised in Table below. The appearance of the lymph nodes was normal in all animals.

Table: DPM, DPN and Stimulation IndexValues for all Groups

 

Test Group Name	Measured DPM / group	DPM	Number of lymph nodes	DPN	Stimulation Index
Background (5% (w/v)TCA)	33.5	-	-	-	-
Negative control (DMF)	4189	4155.5	8	519.4	1.0
HPCTP 10%(w/v) in DMF	2717	2683.5	8	335.4	0.6
HPCTP 5%(w/v) in DMF	4045	4011.5	8	501.4	1.0
HPCTP 2.5%(w/v) in DMF	4226	4192.5	8	524.1	1.0
Positive control(25%(w/v) HCA in DMF)	24456	24422.5	8	3052.8	5.9

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, under the conditions of the present assay, HPCTP, tested in DMF, did not show a sensitisation potential (non-sensitizer) in the Local Lymph Node Assay. No classification/labelling is triggered according to Regulation (EC) No 1272/2008 (CLP) / GHS (rev. 7) 2017.

Executive summary:

The aim of this study was to determine the skin sensitisation potential of HPCTP following dermal exposure in mice. The study was performed with vertebrate animals as classification by use of in vitro alternatives was not possible for this test item. The minimum number of animals was used, corresponding to the regulatory guidelines being followed. The solubility of the test item was examined in a short Preliminary Compatibility Test. The following standard OECD 429 vehicles were assessed: Acetone: Olive oil 4:1 (v/v) mixture, N,N-dimethylformamide (abbreviated as DMF), Methyl ethyl ketone, Propylene glycol, Dimethyl sulfoxide, 1% aqueous Pluronic® PE9200, and cyclohexane. The best vehicle taking into account the test item characteristics, and the requirements of the relevant OECD guideline was considered to be DMF. The 10% (w/v, i.e. 0.1 g per mL with added vehicle) format was the highest concentration which was suitable for the preliminary test. The 10% and 5% (w/v) formulations appeared to be solutions by visual examination. The Preliminary Irritation/Toxicity Test was started according to the Study Plan on CBA/CaCrl mice using two doses (2 animals/dose) with the concentrations of 10% (w/v) and 5% (w/v) in DMF. Based on the results, 10% (w/v) was selected as top dose for the main test.

In the main assay, twenty female CBA/CaCrl mice were allocated to five groups, each group comprised four animals: - three groups of animals received HPCTP (formulated in DMF) at 10, 5, and 2.5% (w/v), respectively, - a negative control group received the vehicle (DMF) only, - a positive control group received 25% (w/v) HCA (dissolved in DMF). The test item solutions were applied to the dorsal surface of the ears of the experimental animals (25 µL/ear) for three consecutive days (Days 1, 2 and 3) and then maintained on study for an additional 3 days. Cell proliferation in the (local) lymph nodes was assessed by measuring disintegrations per minutes after the incorporation of tritiated methyl thymidine ( 3 HTdR) into the lymph nodes and the values obtained were used to calculate stimulation indices (SI) in comparison with the control group. There was no mortality or signs of systemic toxicity observed during the study. No marked body weight losses (≥5%) were observed during the study in any of the animals. The stimulation index values were 0.6, 1.0 and 1.0 at concentrations of 10, 5 and 2.5% (w/v), respectively. The size of lymph nodes was in good correlation with this conclusion. The SI value for the positive control substance α-Hexylcinnamaldehyde (HCA) dissolved in the same vehicle was 5.9, therefore demonstrating the appropriate performance of the assay. In conclusion, under the conditions of the present assay, HPCTP, tested in DMF, did not show a sensitisation potential (non-sensitizer) in the Local Lymph Node Assay. No classification/labelling is triggered according to Regulation (EC) No 1272/2008 (CLP) / GHS (rev. 7) 2017.