Reaction mass of mixed xylenes and sulphur monochloride

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20th Aug 2015 - 8th October 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

22 July 2010

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

no

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:

Reaction Mass of Mixed Xylenes and Sulphur Monochloride

IUPAC Name:

Reaction Mass of Mixed Xylenes and Sulphur Monochloride

Test material form:

liquid

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source:
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 8 to 12 weeks old
- Weight at study initiation:15 to 23g
- Housing: Suspended solid floor polypropylene cages furnished with softwood flakes.
- Diet (e.g. ad libitum): ad libitium
- Water (e.g. ad libitum): ad libitium
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25°C
- Humidity (%): 70%
- Air changes (per hr): 15 per hour
- Photoperiod (hrs dark / hrs light): 12 hours light/ 12 hours dark

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

1,2.5 and 5% v/v

No. of animals per dose:

4

Details on study design:

PRE-SCREEN TESTS
- Irritation: No local skin irritation was observed with undiluted test item or 50, 25, 10 or 5% v/v dilutions.
- Systemic toxicity: The animal dosed with 100% undiluted test item was killed on day 4 due to effects of systemic toxicity. No effects were observed in the surviving animals.
- Ear thickness measurements: Increases in ear thickness were observed in all dose groups by day 6.
50% dose groups : 52.5% increase in ear thickness
25% dose group: 63.6% increase in ear thickness
10% dose group: 27.9% increase in ear thickness
5% dose group: 2.38% increase in ear thickness

MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: The test item will be regarded as as a sensitizer if at least one concentration of the test item results in a thhree fold or greater increase in 3HTdR incorporation compared to control values.

TREATMENT PREPARATION AND ADMINISTRATION:
The mice were treated with a daily application of 25 microlitres of the dose solution to the dorsal surface of each ear for three consecutive days using an automatic micropipette and spread of the surface using the tip.

Positive control substance(s):

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

Results and discussion

Positive control results:

A group of 5 animals was treated wih 50 microlitres (25 per ear) of a 25% v/v solution of alpha-hexylcinnamaldehyde tech (85% purity) in acetone/olive oil 4:1.
A futher control group of 5 animals was dosed with the vehicle.
A stimulation index value of 13.93 was obtained indicating a positive response.

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

ca. 3.11

Variability:

Not applicable - pooled method

Test group / Remarks:

5% v/v

Remarks on result:

other: Positive indication of skin sensitization based on study results.

Results 2

Parameter:

SI

Value:

ca. 0.72

Variability:

Not applicable - pooled method

Test group / Remarks:

2.5% v/v

Remarks on result:

other: Negative indication of skin sensitization based on study results

Results 3

Parameter:

SI

Value:

ca. 0.94

Variability:

Not applicable - pooled method

Test group / Remarks:

1% v/v

Remarks on result:

other: Negative indication of skin sensitization based on study results

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
The proliferation response of lymph node cells was expressed as the number of radioactive disintegrations per minute per lymph node and as a ratio of 3HTdR incorporation into lymph node cells of test nodes relative to that recorded for the control nodes (Stimulation Index).


CLINICAL OBSERVATIONS:
No signs of systemic toxicity were observed for any animal in any dose group over the course of the test or the 3 day observation period after dosing.

BODY WEIGHTS
The body weight change of the dose group animals between day 1 and day 6 was comparable to that of the control group.

Applicant's summary and conclusion

Interpretation of results:

Category 1 (skin sensitising) based on GHS criteria

Conclusions:

The results of the LLNA study (pooled-method) indicated that the test item is a skin sensitizer under the conditions of the test.

Executive summary:

A GLP study was performed in accordance with OECD 429 using the pooled method. Groups of four female mice were dosed with 1, 2.5 and 5% solutions of the test item, reaction mass of mixed xylenes and sulphur monochloride. 25 µl of the dose solution was applied to the dorsal surface of each ear on three consecutive days. Clinical observations and body weights were recorded from days 1 -6. No effects on the body weight or signs of systemic toxicity were observed.

 On day 6 the animals were administered with 3HTdR and 5 hours later the animals were necropsied. The auricular lymph nodes from all animals in each experimental group were excised and pooled.

The Stimulation Index (SI) was calculated for each dose group, an increase of threefold or greater in the 3HTdR incorporation compared to control values was considered a sensitizer. The SI values for the three dose groups 1, 2.5 and 5% were 0.94, 0.72 and 3.11 respectively. The test item is considered to be a sensitizer.