9-Octadecen-1-ol, (Z)-, phosphate

Endpoint summary

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:

2013-04-10 to 2013-04-17

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: according to GLP, OECD and EC guidelines

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

, adopted 22 July 2010

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Version / remarks:

Amendment 06 July 2012 Commission Regulation (EU) No 640/2012

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Version / remarks:

, March 2003

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

other: CBA/Ca mice

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: TOXI-COOP ZRT. H-1103, Budapest, Cserkesz u. 90.
- Age at study initiation: young adult mice, 11-12 weeks (at start of the experiment)
- Weight at study initiation: 17.9 - 21.9 g
- Housing: grouped caging (5 animals/cage)in type II. polypropylene/polycarbonate cages
- Diet: Animals received ssniff® SM R/M-Z+H complete diet for rats and mice produced by ssniff Spezialdiäten GmbH, D-59494 Soest, Germany, ad libitum
- Water: tap water for human supply, ad libitum
-Acclimatization time: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature: :22 ± 3 °C
- Humidity: 30 – 70 %
- Photoperiod: 12 hours daily, from 6.00 a.m. to 6.00 p.m.

Vehicle:

dimethylformamide

Concentration:

5 %, 10 %, 25 %, 50 % and 75 %

No. of animals per dose:

9 animals (1 or 2 animals /group)

Details on study design:

RANGE FINDING TESTS:
- The maximum dose selection was performed according to the relevant guidelines and based on results of the formulation evaluation and the preliminary irritation / toxicity test. The preliminary irritation/toxicity screen was conducted in a similar experimental manner to the exposure phase of the main assay.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay (Individual Approach)
- AOO was used as a vehicle control for the positive control substance. The negative control group for the test item was treated with DMF. The test item was administered at four different concentrations according to the results of the dose range finding test. The positive control substance (HCA) was tested at one concentration.

TREATMENT PREPARATION AND ADMINISTRATION:
Each mouse was topically treated with 25 μL of the appropriate formulations of the test item, of the positive control substance (positive control group) or of the vehicles (AOO or DMF as negative control groups). The formulations were applied, with a pipette, on the dorsal surface of each ear. Each animal was dosed once a day for three consecutive days (Days 1, 2 and 3). There was no treatment on Days 4, 5 and 6.

Clinical Observation:
During the main test (from Day 1 to Day 6) all animals were observed at least once a day for any clinical signs, including systemic toxicity and local irritation. Irritation was monitored by erythema scoring during the whole test (Day 1 to Day 6). Individual records were maintained

Body Weight:
Individual body weights were recorded on Day 1 (beginning of the test) and on Day 6 (prior to 3HTdR injection) with a precision of ± 0.1 g.

Evaluation of the Results:
DPM was measured for each animal. The results were expressed as DPN (DPM divided by the number of lymph nodes). The mean DPM and DPN values were calculated for each treatment group.
The stimulation index (SI = the mean DPN of a treated (positive control or test item) group divided by the DPN of the respective negative control group) for each treatment group was calculated. A stimulation index of 3 or greater is an indication of a positive result. All calculations were made by Microsoft Excel Software. Based on the results the EC3 value (dose calculated to induce a stimulation index of 3) of the test item could not be calculated

Positive control substance(s):

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

Statistics:

Statistical analysis was performed by SPSS/PC+ (4.0.1) software package.
The measured DPM values corrected with the mean background value were used for statistical analysis of the proliferation data. The heterogeneity of variance between groups was checked by Bartlett's test. Since no significant heterogeneity was detected, a one-way analysis of variance was carried out. Inter-group comparisons were performed by Duncan's Multiple Range test. Significance of the positive control response was evaluated by T-test versus the relevant control (AOO).
Significance of the dose-response was evaluated by linear regression made with Microsoft Excel Software.

Positive control results:

The positive control group animals were treated with 25 % (w/v) HCA solution (formulated in AOO) concurrent to the test item groups. No mortality, cutaneous reactions or signs of toxicity were observed in the positive control group.
The positive control substance induced the appropriate, statistically significant stimulation compared to the control (SI = 7.35; p < 0.05, T-test versus AOO control). The results of the positive control item demonstrated appropriate performance of the test in accordance with the relevant guidelines and confirmed sensitivity and validity of the assay.

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

The Stimulation index values per treatment group were as follows: Vehicle control for the positive control AOO: 1.0 SI Vehicle control for the test item DMF: 1.0 SI Positive control: 7.35 SI Test item in DMF: 5.0 %: 2.52 SI 2.5 %: 2.25 SI 1.0 %: 2.43 SI 0.5 %: 1.54 SI

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see Remark

Remarks:

The mean DPM values per treatment group were as follows: Vehicle control for the test item DMF: 736.6 DPM Vehicle control for the positive control AOO: 1299.4 DPM Positive control: 9551.0 DPM Test item in DMF: 5 %: 1858.8 DPM 2.5 %: 1657.8 DPM 1.0 %: 1787.8 DPM 0.5 %: 1135.0 DPM

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of the present Local Lymph Node Assay, Reaction product of oleylalcohol with polyphosphoric acid tested at concentrations of 5 %, 2.5%, 1% and 0.5 % (w/v) as formulations in a suitable vehicle (DMF) was shown to have no skin sensitization potential.

Executive summary:

The aim of this study was to determine the skin sensitization potential of Reaction product of oleylalcohol with polyphosphoric acid in the Local Lymph Node Assay (LLNA). Individual approach was used in this test.

The maximum dose selection was performed according to the relevant guidelines and based on results of the formulation evaluation and a preliminary irritation / toxicity test. Due to significant irritation effect observed in the preliminary test at concentrations of 75 %, 50 %, 25 % and 10 % (w/v) the test item was examined in the main test at concentrations of 5 %, 2.5 %, 1 % and 0.5 % (w/v) as formulations in N,N-Dimethylformamide (DMF).

In the main test, 35 female CBA/Ca mice were allocated to seven groups of five animals each:

- four groups received Reaction product of oleylalcohol with polyphosphoric acid at 5 %, 2.5 %, 1 % or 0.5 % (w/v) in DMF,

- one control group (used as negative control for the test item treated groups) received the vehicle of the test item (DMF) only,

- one control group (used as negative control for the positive control substance treated group) received the vehicle of the positive control substance (AOO) only,

- the positive control group received 25 % (w/v) α-Hexylcinnamaldehyde (HCA) in AOO.

Each substance was applied on the external surface of each ear (25 μL/ear) of the animals for three consecutive days (Day 1, 2 and 3). There was no treatment on Days 4, 5 and 6. On Day 6 animals were intravenously injected via the tail vein with tritiated methyl thymidine (3HTdR), than sacrificed approximately 5 hours after the injection. Auricular lymph nodes were removed and processed. The cell proliferation in the local lymph nodes was measured by incorporation of 3HTdR and the obtained values were used to calculate stimulation indices (SI).

The positive control substance (25 % (w/v) HCA in AOO) induced the appropriate, statistically significant stimulation compared to the relevant control (SI = 7.35; p < 0.05, T-test versus the relevant vehicle (AOO) control), thus confirming the validity of the assay.

No mortality, significant, treatment related effect on the body weights or any other sign of systemic toxicity was observed in any treatment group. No sign of significant irritation (indicated by an erythema score ≥ 3) or any other local effect was observed in any treatment group. Visually larger lymph nodes than the relevant vehicle control (AOO) was observed in the positive control group. Visual appearance of the lymph nodes was normal in the vehicle (both AOO and DMF) control groups and in the test item treated groups.

No significantly increased lymphoproliferation (≥ 3) was observed for the test item at the tested concentrations. The observed SI values were 2.52, 2.25, 2.43 and 1.54 at treatment concentrations of 5 %, 2.5 %, 1 % and 0.5 % (w/v), respectively. No statistically significant differences compared to the relevant vehicle (DMF) control were observed. No significant dose-related response was observed (linear regression, p = 0.35, r = 0.65).

Since the test was valid and no sign of systemic toxicity or significant irritation was observed, the proliferation values obtained are considered to reflect the real potential of the test item to cause/not cause lymphoproliferation in the Local Lymph Node Assay.

According to evaluation criteria of the relevant guidelines [1-3], the lack of a significantly increased proliferation value up to the maximum applicable concentrations of 5 % (w/v) and the lack of a dose-related response are considered as evidence that Reaction product of oleylalcohol with polyphosphoric acid is not a skin sensitizer.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

The aim of this study was to determine the skin sensitization potential of Reaction product of oleylalcohol with polyphosphoric acid in the Local Lymph Node Assay (LLNA). Individual approach was used in this test.

The maximum dose selection was performed according to the relevant guidelines and based on results of the formulation evaluation and a preliminary irritation / toxicity test. Due to significant irritation effect observed in the preliminary test at concentrations of 75 %, 50 %, 25 % and 10 % (w/v) the test item was examined in the main test at concentrations of 5 %, 2.5 %, 1 % and 0.5 % (w/v) as formulations in N,N-Dimethylformamide (DMF).

In the main test, 35 female CBA/Ca mice were allocated to seven groups of five animals each:

- four groups received Reaction product of oleylalcohol with polyphosphoric acid at 5 %, 2.5 %, 1 % or 0.5 % (w/v) in DMF,

- one control group (used as negative control for the test item treated groups) received the vehicle of the test item (DMF) only,

- one control group (used as negative control for the positive control substance treated group) received the vehicle of the positive control substance (AOO) only,

- the positive control group received 25 % (w/v) α-Hexylcinnamaldehyde (HCA) in AOO.

Each substance was applied on the external surface of each ear (25 μL/ear) of the animals for three consecutive days (Day 1, 2 and 3). There was no treatment on Days 4, 5 and 6. On Day 6 animals were intravenously injected via the tail vein with tritiated methyl thymidine (3HTdR), than sacrificed approximately 5 hours after the injection. Auricular lymph nodes were removed and processed. The cell proliferation in the local lymph nodes was measured by incorporation of 3HTdR and the obtained values were used to calculate stimulation indices (SI).

The positive control substance (25 % (w/v) HCA in AOO) induced the appropriate, statistically significant stimulation compared to the relevant control (SI = 7.35; p < 0.05, T-test versus the relevant vehicle (AOO) control), thus confirming the validity of the assay.

No mortality, significant, treatment related effect on the body weights or any other sign of systemic toxicity was observed in any treatment group. No sign of significant irritation (indicated by an erythema score ≥ 3) or any other local effect was observed in any treatment group. Visually larger lymph nodes than the relevant vehicle control (AOO) was observed in the positive control group. Visual appearance of the lymph nodes was normal in the vehicle (both AOO and DMF) control groups and in the test item treated groups.

No significantly increased lymphoproliferation (≥ 3) was observed for the test item at the tested concentrations. The observed SI values were 2.52, 2.25, 2.43 and 1.54 at treatment concentrations of 5 %, 2.5 %, 1 % and 0.5 % (w/v), respectively. No statistically significant differences compared to the relevant vehicle (DMF) control were observed. No significant dose-related response was observed (linear regression, p = 0.35, r = 0.65).

Since the test was valid and no sign of systemic toxicity or significant irritation was observed, the proliferation values obtained are considered to reflect the real potential of the test item to cause/not cause lymphoproliferation in the Local Lymph Node Assay.

According to evaluation criteria of the relevant guidelines [1-3], the lack of a significantly increased proliferation value up to the maximum applicable concentrations of 5 % (w/v) and the lack of a dose-related response are considered as evidence that Reaction product of oleylalcohol with polyphosphoric acid is not a skin sensitizer.

Migrated from Short description of key information:
Under the conditions of the present Local Lymph Node Assay, Reaction product of oleylalcohol with polyphosphoric acid tested at concentrations of 5 %, 2.5%, 1% and 0.5 % (w/v) as formulations in a suitable vehicle (DMF) was shown to have no skin sensitization potential.

Justification for selection of skin sensitisation endpoint:
Most reliable study.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the results obtained Reaction product of oleylalcohol with polyphosphoric acid has no sensitisation potential and was not classified and labeled according to Regulation (EC) No 1272/2008 (CLP) and Directive 67/548/EEC (DSD).