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Toxicological information

Skin sensitisation

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Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
27 February 2007 to 6 March 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
The test substance was identified by name only. No structural characterization or purity information was available.
Cross-reference
Reason / purpose:
read-across: supporting information
Reference

Skin sensitisation (OECD TG 429): Not skin sensitising (read-across from Dihymyrcetol) and absence of electrophilic groups

Respiratory sensitisation: Not respiratory sensitising in view of absence such data on humans and because of absence of skin sensitisation.

Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

The skin sensitisation of Tetrahydromyrcenol is assessed by using read across from Dimyrcetol. First the experimental skin sensitisation information of Dimyrcetol will be summarised. Thereafter the read across justification is presented, the accompanying files are attached in the present Sensitisation Endpoint summary.

Skin sensitisation information of Dimyrcetol

Dimyrcetol was assessed for skin sensitizing potential using the mouse Local Lymph Node Assay (OECD TG 429). The material was applied as 0.5, 1, 2.5, 10 or 25% w/v preparations in 1:3 ethanol:diethylphthalate. The test material failed to produce a 3 -fold increase in lymphocyte proliferation and the EC3 value was estimated to be in excess of 6250 micrograms/cm2. Based on the results of this study it is concluded that the test substance Dimyrcetol does not need to be classified as a potential skin sensitizer.

Tetrahydromyrcenol (Cas no18479-57-7)and its non-skin sensitising properties using read across from Dimyrcetol (mixture of Dihydromyrcenol and its formate ester (25279-09-8)

Introduction and hypothesis for the read across

Tetrahydromyrcenol is a branched C8 alkyl chain to which a tertiary alcohol is attached. For this substance noskin sensitisation data are available. Therefore additional information is used in accordance with Article 13 of REACH where it is said thatlacking information should be generated whenever possible by means other than vertebrate animal tests, i.e. applying alternative methods such as in vitro tests, SARs, grouping and read-across.

Hypothesis:Tetrahydromyrcenolis not expected to have sensitising properties based on skin sensitisation information from Dimyrcetol.

Available information: For Dimyrcetol an LLNA is available (OECD TG 429) in which the maximum concentration used is 25%, with absence of skin sensitisation.

Target and Source chemical(s):

Chemical structures of the Tetrahydromyrcenol (the target) and are shown in the Data matrix including physico-chemical properties and toxicological information, thought relevant for skin sensitisation.

Purity / Impurities:

The purity and impurities of the target chemical do not indicate skin sensitisation potential other than indicated by the parent substance. The impurities are all below 10%.

Analogue justification

According REACH Annex XI an analogue approach and structural alert information can be used to replace testing when information from different sources provides sufficient evidence to conclude that this substance has or does not have a particular dangerous property. The result derived should be applicable for C&L and/or risk assessment and be presented with adequate and reliable documentation.

Analogue selection: Dihydromyrcenol /Dihydromycenyl formate and2,6-Dimethyl-2-heptanol (Cas no. 13254-34-7)were considered as potential analogues. When using the Tanimoto index (MSC) the similarity with Dihydromyrcenol was > 80%. Dihydromyrcenol / Dihydromycenyl formate was considered the more conservative analogue having an electrophilic double bond at one end of the alkyl chain.

Structural similarities and differences:The target Tetrahydromyrcenol and the source chemical Dihydromyrcenol have a 2,6-dimethyl)octan-2-ol backbone and both have a tertiary alcohol functional group at one end of the alkyl chain. The difference is that the alkyl chain of Tetrahydromyrcenol is fully saturated, while Dihydromyrcenol has a double bond at the end of the alkyl chain. The formate ester of Dihydromyrcenol has a formic acid group at the tertiairy alcohol site.

Toxico-kinetics: To some extent skin absorption is needed for skin sensitisation. Tetrahydro- and Dihydromyrcenol /formate are likely to absorb in skin. They are both liquids have molecular weights of 158 and 156/184, respectively. Also the physico-chemical properties present skin absorption potential equally for both substances. Skin metabolism is likely for the formate ester by skin carboxylase and will become Dihydromyrcenol and formic acid. Oxidation of the double bond may also occur.

 

 

 

 

Fig. 1    The two components of Dimyrcetol are shown including the sites where the substance can metabolise

 

Toxico-dynamics: Tetrahydromyrcenol has no electrophilic features that present electrophilicity which is a key parameter to cause skin sensitisation. In view of being fully saturated it does not have oxidation-site. Dihydromyrcenol has this double bond at one end of the alkyl chain and at that position is slightly more reactive, because potentially it may oxidise. In absence of a methyl group that stabilised an oxidation product, therefore no skin sensitisation activity is anticipated. If anything Dihydromyrcenol /formate would be more prone to skin sensitisation activity based on this double, more electrophilic, bond.

Other toxicological information present for supporting the read across: Tetrahydro- and Dihydromyrcenol are both skin and eye irritants, which indicate similar reactivity.

Remaining uncertainties:It is presented above that Dihydromyrcenol/formate can be considered a conservative approach. It is noted that the analogue is tested up to 25%, which may be the highest dose in view of skin irritant properties. In absence of reactive groups and absence of pre or pro-hapten features it can be concluded that Tetrahydromyrcenol will not be sensitising up to 100%.

Conclusions on the skin sensitisationper endpoint

For Tetrahydromyrcenol the skin sensitisation potential can be derived from Dimyrcetol using read across which was negative in the LLNA up to 25% (OECD TG 429). In addition, Tetrahydromyrcenol has no structural features that indicate potential skin sensitisation and therefore absence of skin sensitisation is anticipated up to 100%.

Final conclusion on hazard and C&L: Tetrahydromyrcenol is not a skin sensitiser based on analogue information from Dihydromyrcenol and absence of sensitisation structural features.

Data matrix: Tetrahydromyrcenol and its analogue Dihydromyrcenol.

Common names

Tetrahydromyrcenol

Dihydromyrcenol

Chemical structures

CAS no

18479-57-7

18479-58-8

REACH registration

For 2018

2010 registered.

Tanimoto *

1

0.83

Molecular weight

158.29

156.27

Physico-chemical data

 

 

Physical state

Liquid

Liquid

Melting point,oC

<-20

<-20 (ECHA dissemination site)

Boiling point,oC

192.80

193 (ECHA dissemination site)

Vapour pressure, Pa

9.3

20 (ECHA dissemination site)

Water solubility, mg/l

281.9

939 (ECHA dissemination site)

Log Kow

3.2 (m)

3.25 (ECHA dissemination site)

Human health endpoints

 

 

Skin sensitisation

(Read across from Dihydromyrcenol

Not sensitizing

 

Genotoxicity – Ames test

Not mutagenic

(OECD TG 471)

Not mutagenic

(OECD TG 471)

 

Tanimoto calculation:http://chemmine.ucr.edu/similarity/#

 

Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Respiratory sensitisation can be assessed using human data such as indicated in R7.3.5.2 of the ECHA guidance (2015) that indicate respiratory reactions e. g. from consumer experience or occupational exposure. In case no such data are available the respiratory sensitisation can be assessed using the integrated evaluation strategy for respiratory sensitisation data in the ECHA guidance (R7A, Fig. 7.3-2, 2015), which says that if the substance is not a skin sensitiser, it is unlikely to be a respiratory sensitiser.

Based on the available data, it can be concluded that Tetrahydromyrcenol does not need to be classified as a skin sensitiser and is therefore also not a respiratory sensitiser in accordance with the criteria in EU CLP (EC 1272/2008 and its updates).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Test material form:
liquid

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/Ca
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River UK Ltd., Manston Road, Margate, Kent, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 16.9 to 22.3 g
- Housing: A maximum of 4 mice/cage
- Diet (ad libitum): RM1, Special Diet Services Ltd., Witham, Essex, UK
- Water (ad libitum): mains
- Acclimation period: At least 5 days prior to start of dosing


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30-70
- Air changes (per hr): Minimum of 15
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: 27 February 2007 To: 6 March 2007

Study design: in vivo (LLNA)

Vehicle:
other: 1:3 Ethanol:Diethylphthalate (CTL Ref: Y05722/002)
Concentration:
0.5, 1, 2.5, 10 or 25% w/v
No. of animals per dose:
4
Details on study design:
Dose Preparation:
All dose preparations were used within 24 hours. Stability and achieved concentrations were not determined.

Test Method Details:
Groups of 4 mice were used at each dose level. Approximately 25 microliters of 0.5, 1, 2.5, 10 or 25% w/v preparations of the test substance in vehicle were applied to the dorsal surface of each ear. A vehicle control group was similarly treated with vehicle alone. This procedure was repeated daily for 3 consecutive days.

Three days after the last application, all animals were injected with approx. 250 microliters of phosphate buffered saline containing 20 microCuries of a 2.0 Ci/mmol specific activity of 3H-methyl thymidine. Approximately 5 hours later, the animals were humanely killed by inhalation of halothane vapour followed by cervical dislocation. The draining auricular lymph nodes were removed from each animal and, together with the nodes from the other animals in the group, were placed in a container of PBS.

A single cell suspension was prepared by mechanical disaggregation of lymph nodes through a 200-mesh stainless steel gauze. The cell suspensions were then washed three times by centrifugation with approximately 10 ml of PBS. Approximately 3 ml of 5% w/v trichloroacetic acid (TCA) was added and, after overnight precipitation at 4 deg C, the samples were pelleted by centrifugation and the supernatant was discarded. The cells were resuspended in approx. 1 ml of TCA.

The lymph node suspensions were transferred to scintillation vials and 10 ml of scintillant (Optiphase) was added prior to beta-scintillation counting using a Packard Tri-Carb 3100TR Liquid Scintillation Counter.

Clinical Observations:
Animals were checked at least once daily for signs of systemic toxicity. Animals were monitored for any signs of irritancy on the ears on days 1, 2, 3 and 6 of study. Irritancy was scored at a visual level with the scoring scheme of none detectable, mild, moderate or severe.

Body Weights:
The body weight of each animal was recorded prior to dosing on day 1 and prior to injection of the 3H-methyl thymidine on day 6.

Positive Control:
Approximately 25 microliters of 5%, 10% or 25% of the positive control in acetone in olive oil (4:1) was applied, and a vehicle control of acetone:olive oil was similarly treated.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
None described

Results and discussion

Positive control results:
The application of the positive control substance at the 25% w/v concentration resulted in greater than a 3-fold increase in isotope incorporation.

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
other: EC3 was not reached
Remarks on result:
other: At 25% the 3-fold increase in lymphocyte proliferation (EC3) was not reached.
Parameter:
SI
Value:
0.9
Remarks on result:
other: 0.5%
Parameter:
SI
Value:
0.8
Remarks on result:
other: 1%
Parameter:
SI
Value:
0.8
Remarks on result:
other: 2.5%
Parameter:
SI
Value:
0.9
Remarks on result:
other: 10%
Value:
1.4
Remarks on result:
other: 25%

Any other information on results incl. tables

Table:  Skin Sensitization Results for LLNA-965

Concentration of test substance (%w/v)

Number of lymph nodes assayed

Disintegrations per minute (dpm)

dpm per lymph node

Test: Control ratio (SI)

0 (vehicle only)

8

4857

607

N/A

0.5

8

4435

554

0.9

1

8

3646

456

0.8

2.5

8

4053

507

0.8

10

8

4236

530

0.9

25

8

6565

821

1.4

Applicant's summary and conclusion

Interpretation of results:
other: not skin sensitising
Remarks:
based on EU CLP criteria (1272/2008/EC and its updates))
Conclusions:
Under the conditions of this test, it can be concluded that the test substance Dimyrcetol does not need to be classified as a skin sensitizer in accordance with the criteria outlined in Annex I of the CLP Regulation (1272/2008/EC).
Executive summary:

Dimyrcetol was assessed for skin sensitizing potential using the mouse Local Lymph Node Assay (OECD TG 429). The material was applied as 0.5, 1, 2.5, 10 or 25% w/v preparations in 1:3 ethanol:diethylphthalate. The test material failed to produce a 3 -fold increase in lymphocyte proliferation and the EC3 value was estimated to be in excess of 6250 micrograms/cm2. Based on the results of this study it is concluded that the test substance Dimyrcetol does not need to be classified as a potential skin sensitizer.