Propanoic acid, 2-[4-[4,6-bis([1,1'-biphenyl]-4-yl)-1,3,5-triazin-2-yl]-3-hydroxyphenoxy]-, isooctyl ester

Endpoint summary

Description of key information

A local lymph node assay in mice was conducted to assess the allergenic potential of the test material (according OECD guideline 429, GLP). Mice were treated by daily application of 25 µl of the respective test material (10 - 50 % solution in 4:1 v/v acetone/olive oil) to the dorsal surface of both ears for three consecutive days. A a 3-fold greater increase in 3HTdR incorporation was not observed. The test item is not regarded as a potential skin sensitizer.

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:

29-Oct-2002 to 12-Nov-2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study (OECD test guideline 429)

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

24th April 2002

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan UK Ltd., Bicester, Oxon, England
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 15.9-20.8 g
- Housing: The mice were allocated without conscious bias to cages within the treatment groups. They were housed individually in polycarbonate cages with woodflake bedding.
- Diet: standard laboratory rodent diet (ad libitum)
- Water: drinking water (ad libitum)

ENVIRONMENTAL CONDITIONS
- Temperature: 21+/- 3 °C
- Humidity: 40-70%
- Photoperiod: 12 hours dark / 12 hours light

IN-LIFE DATES: From: 31-Oct-2002, necrospy date not indicated

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

10, 25 and 50% (w/v); the maximum practical concentration of the test substance for pinna dosing was 50% w/v in acetone:olive oil (4:1 v/v). Based on this information the concentrations were selected.

No. of animals per dose:

5 mice

Details on study design:

TREATMENT PREPARATION AND ADMINISTRATION:
TOPICAL APPLICATION:
The mice were treated by daily application of 25 µL of the appropriate concentration of the test substance to the dorsal surface of each ear for three consecutive days (Days 1-3). The test substance was applied to the dorsal surface of each ear using an automatic micropipette. The test substance was spread over the entire dorsal surface of the ear using the tip of the pipette. In addition, a group of five mice was similarly dosed with the vehicle alone and a further group of five mice received HCA (positive control) in the same manner.
ADMINISTRATION OF 3H-METHYL THYMIDINE:
Five days following the first topical application of test substance (Day 6) all mice were injected via the tail vein with 250 µL of physiological saline containing 3H-methyl thymidine (3-HTdR: 80 µCi/mL) giving a nominal of 20 µCi to each mouse. The injection into the tail vein was carried out using a plastic syringe and needle (26 gauge) with the mouse restrained in a hot box.
TERMINATION:
Five hours following the administration of 3-HTdR on Day 6 all mice were killed by carbon dioxide asphyxiation and the draining auricular lymph nodes excised. 1.0 mL of physiological saline was added to the lymph nodes for each animal. No further investigations were carried out.
PREPARATION OF SINGLE CELL SUSPENSIONS:
A single cell suspension of lymph node cells (LNC) for each individual animal was prepared by gentle mechanical disaggregation through a stainless steel gauze (200 mesh size). The LNC were then washed by adding 10 ml physiological saline, pelleted at 190 x g for 10 minutes and resuspended. The cells were washed twice again and resuspended in 3 ml trichloroacetic acid (TCA: 5%) following the final wash.
DETERMINATION OF INCORPORATED 3H-METHYL THYMIDINE:
After overnight incubation (at least 18 hours) with 5% TCA at 4°C, the precipitate was recovered by centrifugation (10 minutes at 190 x g) and resuspended in 1 mL 5% TCA and transferred to 10 mL Ultima Gold scintillation fluid on Day 7. 3-HTdR incorporation was measured by P-scintillation counting. The proliferative response of LNC was expressed as radioactive disintegrations per minute per lymph node (dpm/node) and as the ratio of 3-HTdR incorporation into LNC of test nodes relative to that recorded for vehicle control nodes (test/control ratio).

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated in the body weight tables.

Positive control results:

Greasy fur in the cervical region was noted for all animals post-dose from Day 1 and prior to dosing from Day 2. This sign was also noted on Days 4 to 6. Stimulation index for the positive control substance hexyl cinnamic aldehyde (HCA) was 6.7 at concentrations of 25%, which proved to demonstrate the reliability and sensitivity of this assay to detect skin sensitization potential.

Parameter:

other: disintegrations per minute (dpm) / node

Value:

1 575

Test group / Remarks:

Positive control (hexyl cinnamic aldehyde, 25% v/v)

Parameter:

other: disintegrations per minute (dpm) / node

Value:

236

Test group / Remarks:

Vehicle control (acetone:olive oil, 4:1 v/v)

Parameter:

other: disintegrations per minute (dpm) / node

Value:

175

Test group / Remarks:

Test group (10% w/v)

Parameter:

other: disintegrations per minute (dpm) / node

Value:

97

Test group / Remarks:

Test group (25% w/v)

Parameter:

other: disintegrations per minute (dpm) / node

Value:

422

Test group / Remarks:

Test group (50% w/v)

Parameter:

SI

Value:

6.7

Test group / Remarks:

Positive control (hexyl cinnamic aldehyde, 25% v/v)

Parameter:

SI

Test group / Remarks:

Vehicle control (acetone:olive oil, 4:1 v/v)

Remarks on result:

other: not applicable

Key result

Parameter:

SI

Value:

0.7

Test group / Remarks:

Test group (10% w/v)

Key result

Parameter:

SI

Value:

0.4

Test group / Remarks:

Test group (25% w/v)

Key result

Parameter:

SI

Value:

1.8

Test group / Remarks:

Test group (50% w/v)

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
The stimulation indices obtained for 10, 25 and 50% w/v test substance were 0.7, 0.4 and 1.8
respectively. As a stimulation index of 3 or more was not recorded for any of the concentrations tested, the test substance is not considered to have the potential to cause skin sensitization (delayed contact hypersensitivity).

DETAILS ON STIMULATION INDEX CALCULATION
Results for each treatment group were expressed as the stimulation index (SI). This was obtained by comparing the proliferation in the vehicle treated control group with the values from the three test groups as follows: the ratio of 3HTdR incorporation into lymph node cells, expressed as dpm, relative to that recorded for confrol lymph nodes is derived for each test group based on the group mean dpm per node. When the stimulation index >3, the test substance is regarded as a skin sensitizer with a consideration given to dose response and statistical significance.

CLINICAL OBSERVATIONS AND SIGNS OF TOXICITY (including dermal irritation at the site of administration, if any, e.g. increased ear thickness).
- There were no deaths and no signs of ill health or toxicity observed during the study.
- Greasy fur in the cervical region was noted post-dose and prior to dosing for all animals of the test and control groups, respectively.
- White particles on the ears were noted for three females of test group 25% w/v.
- Hairloss, reddening of the skin on the head and white particles on the ears were noted for all animals of test group 50% w/v.

BODY WEIGHTS
Please, see table 1 in section 'Any other information on result incl. tables'

Table 1: Individual and group mean bodyweights and individual bodyweight changes (g)

 

Group	Animal number	Day 1*	Termination (Day 6)	Individual bodyweight changes (Day 1 to Day 6)
1 Vehicle control	B1	16.3	17.8	1.5
	B2	20.3	22.4	2.1
	B3	18.1	20.7	2.6
	B4	18.1	19.0	0.9
	B5	19.8	20.3	0.5
	Mean	18.5	20.0	-
2	B6	19.3	21.8	2.5
	B7	16.9	18.9	2.0
	B8	17.7	18.5	0.8
	B9	17.9	20.1	2.2
	B10	20.5	21.4	0.9
	Mean	18.5	20.1	-
3	B11	17.1	19.3	2.2
	B12	19.5	20.7	1.2
	B13	20.5	23.4	2.9
	B14	19.7	21.7	2.0
	B15	20.8	21.1	0.3
	Mean	19.5	21.2	-
4	B16	18.6	19.8	1.2
	B17	20.3	22.2	1.9
	B18	20.0	20.2	0.2
	B19	19.4	21.5	2.1
	B20	18.1	20.1	2.0
	Mean	19.3	20.8	-
5 Positive control	B21	19.1	20.6	1.5
	B22	19.3	21.5	2.2
	B23	19.4	22.0	2.6
	B24	17.5	19.2	1.7
	B25	15.9	17.1	1.2
	Mean	18.2	20.1	-

- Not applicable

* Prior to dosing

Interpretation of results:

GHS criteria not met

Conclusions:

The test item is not regarded as a potential skin sensitizer.

Executive summary:

A local lymph node assay in mice was conducted to assess the allergenic potential of the test material (according OECD guideline 429 and GLP). Mice were treated by daily application of 25 µl of the respective test (10 - 50 % solution in 4:1 v/v acetone/olive oil) or control (negative and positive) material to the dorsal surface of both ears for three consecutive days. The proliferative response of the lymph node cells (LNC) from the draining auricular lymph nodes was assessed five days following the initial application, by measurement of the incorporation of 3H-Thymidine by beta-scintillation counting of LNC suspensions. The response was expressed as radioactive disintegrations per minute per lymph node (dpm/node) and as the ratio of 3HTdR incorporation into LNC of test nodes relative to that recorded for control nodes (test/control ratio).

In this assay, a 3-fold greater increase in 3HTdR incorporation compared to the vehicle control values was not observed. As a result, the test item is not regarded as a potential skin sensitizer.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Procedure and observation

A local lymph node assay in mice was conducted to assess the allergenic potential of the test material. Therefore, each group of mice was treated by daily application of 25 µl of the respective test (10 - 50 % solution in 4:1 v/v acetone/olive oil) or control (negative or positive) material, to the dorsal surface of both ears for three consecutive days. The proliferative response of the lymph node cells (LNC) from the draining auricular lymph nodes was assessed five days following the initial application, by measurement of the incorporation of 3H-Thymidine by beta-scintillation counting of LNC suspensions. The response was expressed as radioactive disintegrations per minute per lymph node (dpm/node) and as the ratio of 3HTdR incorporation into LNC of test nodes relative to that recorded for control nodes (test/control ratio).

The test substance is regarded as a sensitizer if at least one concentration of the chemical results in a 3-fold greater increase in 3HTdR incorporation compared to the vehicle control values. In this assay the test/control ratios obtained for 10, 25 and 50% w/v were 0.7, 0.4 and 1.8 respectively which indicates that the substance did not show the potential to induce skin sensitization (delayed contact hypersensitivity).

 

Conclusion

The test item is not regarded as a potential skin sensitizer.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. The stimulation indices in the LLNA (OECD 429) did not show a dose dependent increase. Therefore, the substance does not require classification as a skin sensitizer under Regulation (EC) No. 1272/2008, as amended for the fourteenth time in Regulation (EC) No. 2020/217.