Linalool

Endpoint summary

Description of key information

In a local lymph node assay according to OECD guideline 429, the test item was found to be a skin sensitiser and an EC3 value of 35.5 % (w/v) was derived.

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study performed according to OECD 429 and GLP

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Envigo RMS B.V., Inc
- Age at study initiation:
Pre-test: 10 - 11 weeks
Main study: 9 - 10 weeks
- Weight at study initiation: 18.6 - 21.2 g
- Assigned to test groups randomly: Yes
- Housing: Animals were distributed into the test groups at random, all animals belonging to the same experimental group were kept in one cage
- Diet: 2018C Teklad Global 18 % protein rodent diet (certified), ad libitum
- Water: Tap water ad libitum
- Acclimation period: At least 5 days prior to the start of dosing under test conditions after health examination

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 45 - 65
- Photoperiod (hrs dark / hrs light): 12 / 12

Vehicle:

dimethylformamide

Concentration:

25, 50 and 100%

No. of animals per dose:

5

Details on study design:

Three groups each of five female mice were treated with different concentrations of the test item by topical application at the dorsum of each ear once daily each on three consecutive days. A control group of five mice was treated with the vehicle only. Five days after the first topical application, the mice were intravenously injected into a tail vein with radio-labelled thymidine (3H-methyl thymidine; 3HTdR). Approximately five hours after intravenous injection, the mice were sacrificed and the draining auricular lymph nodes were excised and pooled per animal. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes, which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells were then determined by the incorporation of 3H-methyl thymidine measured in a β-scintillation counter.

Positive control substance(s):

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

Statistics:

A statistical analysis was conducted on the ear weights to assess whether the difference was statistically significant between test item groups and
negative control (vehicle) group. For all statistical calculations validated statistical program R Script DecisionTree_2.Rnw was used.
Statistical significance was set at the five per cent level (p < 0.05).

Positive control results:

The sensitivity and reliability of the experimental technique employed was assessed by use of α-hexyl cinnamaldehyde dissolved in acetone/olive oil (4+1 v/v) (compound listed in OECD 429 Guideline) which is known to have skin sensitisation properties in mice. The periodic positive
control experiment was performed using CBA/CaOlaHsd mice in October 2015.
Result: - 10 % α-Hexylcinnamaldehyde (in acetone/olive oil, 4+1 v/v): S.I. 4.23
- 25 % α-Hexylcinnamaldehyde: S.I. 17.56

Key result

Parameter:

SI

Value:

1

Test group / Remarks:

5 animals/ 25%

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: - vehicle control group (DMF): mean DPM per animal: 835.5, SD 335.78 - 25% Linalool: mean DPM per animal: 2073.9, SD 433.09 - 50% Linalool: mean DPM per animal: 3104.5, SD 814.95 - 100% Linalool: mean DPM per animal: 3564.3, SD 1086.55

Key result

Parameter:

SI

Value:

2.48

Test group / Remarks:

5 animals/50%

Key result

Parameter:

SI

Value:

3.72

Test group / Remarks:

5 animals / 100%

EC3 = 35.5% (w/v)

Interpretation of results:

Category 1B (indication of skin sensitising potential) based on GHS criteria

Conclusions:

The test item Linalool was found to be a skin sensitiser and an EC3 value of 35.5 % (w/v) was derived.

Executive summary:

In the study the test item Linalool formulated in water-free dimethylformamide (DMF) was assessed for its possible skin sensitising potential. For this purpose a local lymph node assay was performed using test item concentrations of 25, 50, (w/v) and 100%.

No cases of mortality were observed. From day 1 to 4, the animals showed an erythema of the ear skin (Score 1, animals treated with 25%: 1h after the third application, animals treated with 50%: 1h after the second and third application, animals treated with 100%: 1h after the first and second application, on days 3 and 4). Transiently, especially after applications, nervousness, tumbling and burrowing in the bedding were observed in some of the animals (for details see Appendix 3). A relevant increase in ear weights was not observed in any of the treated groups in comparison to the vehicle control group.

In this study Stimulation Indices (S.I.) of 2.48, 3.72, and 4.27 were determined with the test item at concentrations of 25, 50, and 100%, respectively.

The test item Linalool was found to be a skin sensitiser and an EC3 value of 35.5 % (w/v) was derived. The substance has to be classified according to criteria outlined in Annex I of the Regulation (EC) No. 1272/2008.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

Skin sensitisation, mouse, RL1

A Local Lymph Node Assay (LLNA, Remus & Dony, 2016) was performed using three groups of female mice (5 mice per group). The animals were treated with different concentrations of linalool (25, 50 (w/v) and 100%) by topical application at the dorsum of each ear once daily on three consecutive days. A control group of five mice was treated with the vehicle (DMF) only. Five days after the first application, the mice were intravenously injected into a tail vein with radiolabelled thymidine, then sacrificed and the lymph nodes were pooled per animal. No relevant increase in ear weights were observed and no relevant erythema was observed. In this study, Stimulation Indicis (S.I.) of 2.48, 3.72 and 4.27 were determined with the test item at concentrations of 25, 50 and 100%, respectively. Linalool was found to be a skin sensitizer and an EC3 value of 35.5% (w/v) was derived.

 

Supporting Human Information: A Human Repeated Insult Patch Test (HRIPT) on 135 healthy volunteers did not show skin sensitization potential of linalool (Harrison & Spey 2005). The skin of 45 male and 90 female was repeatedly exposed (induction, rest, challenge phase) to three patches of which one was containing 12.7% linalool. Saline and the vehicle (1:3 EtOH:DEP) were used as controls. Both, erythema and edema were assessed and scored after every application period and 24, 48 and 72 hours after removal of the patches in the challenge phase. 119 human subjects completed the study (36 male and 83 female). The test revealed that linalool induced slight erythema in 1 out of 117 and 119 human subjects at first and second reading after challenge.

The substance is a skin sensitizer (EC3 = 35.5% w/v).

 

Skin sensitisation, human, RL1 (please refer to section 7.10.4)

The senstiziting potential of 12.7% linalool was tested in this human Repeated Insult Patch Test (RIPT). Saline and the vehicle (1:3 EtOH:DEP) were used as control. The skin of 135 human subjects, 45 male and 90 female, was repeatedly exposed (induction, rest, challenge phase) to three patches containing all three substances. During induction, 9 applications of the substance were done in three weeks. Following a rest period of 2 weeks, a challenge was done by application of the same substance on the other side (previously untreated) of the back. Erythema and also edema were assessed and scored after every application period (induction and challenge). In the challenge phase, reactions were scored at 0, 24, 48 and 72 hours after removal of patch with test substance. 119 subjects completed the study (36 male and 83 female). During induction phase 6 reactions were noted for saline, 4 reactions for linalool, and 2 reactions for the vehicle. All reactions were faint, minimal erythema. Linalool only induces slight erythema in one out of 117 and 119 subjects assessed at second and third reading after challenge. 5 reactions after challenge (faint, minimal erythema to erythema) were seen in the saline group. 2 reactions (faint, minimal erythema) were observed in the vehicle group. No difference in effects was observed between application of the test substance and the control substances. When considering the criteria outlined in Annex I of Regulation (EC) No. 1272/2008, linalool will not have to be classified as a sensitiser up to a concentration of 12.7%.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the available information, linalool is considered as skin sensitiser. Therefore, the substance linalool needs to be classified as skin sensitizer Cat. 1B according to Regulation (EC) No 1272/2008, as amended for fifteenth time in Regulation (EU) No 2020/1182.