Dodecane-1,12-diol

Endpoint summary

Description of key information

Due to the inconsistency of the results in silico (no alert), in chemico (negative) and in vitro KeratinoSens (positive) no final conclusion for skin sensitization can be drawn. Thus, the mouse LLNA was initiated. The registered substance did not reveal any skin sensitising properties in the LLNA.

In conclusion, no skin sensitizing effects were observed in vivo.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Reference 1

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

- modified LLNA = Local Lymph Node Assay (LLNA) - Integrated Model for Differentiation of Skin reactions (IMDS)): Measurement of cell counts instead of radioactive labeling. In addition, measurements of ear swelling and ear weights were done to discrimina

Principles of method if other than guideline:

Modified LLNA (IMDS; Integrated Model for the Differentiation of Skin Reactions). Modifications are authorized in the OECD TG 429 and in the Note for Guidance SWP/2145/00 of the CPMP (2001). Information on validation of IMDS and scientific justification is given in: Vohr HW et al., Arch. Toxicol., 73, 501-509 (2000); Ehling G et al., Toxicology 212, 60-68 and 69-79 (2005).
The study was performed according to OECD 429, however not employing the use of radioactive labelling to measure cell proliferation, as the radioactive method proposed by the OECD guideline led to problems in various EU laboratories: such as (i) practical difficulties/complexity of the test, in particular the radiochemical steps, which sometimes resulted in loss of specimen/activity; this in turn led to variability in the results and to a poor reproducibility and (ii) radiation protection issues. However, the OECD guideline allows other endpoints for assessment of proliferation in form of lymph node cell counts and lymph node weights if justification and appropriate scientific support exist showing the validity of this method.
The alternative method used for the study employing the lymph node weight and lymph node cell count to assess proliferation has been established by an European interlaboratory validation exercise, as described in the two publications by Ehling et al. 2005a and 2005b. This method has the advantage of (i) more simplistic experimental work, (ii) less variability, (iii) better reproducibility, (iv) faster results, (v) reduced costs.
In addition, the acute inflammatory skin reaction is measured by ear weight determination of circular biopsies of the ears and ear thickness measurements on test day 1 and test day 4 to identify skin irritation properties of the test item.

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: Due to the limited solubility, the test item was dissolved in dimethyl sulfoxide (DMSO) and heated up to 37 °C to gain the maximal feasible concentration of 10% (w/w).
DMSO was selected as vehicle as it provided a suitable solution of the test item both for administration and adherence to the mouse ear and is recommended by the OECD guideline. Acetone/olive oil (4:1, v/v), methyl ethyl ketone, propylene glycol and N,N-dimethylformamide, other recommended vehicles, did not provide higher concentrated suitable solutions or suspensions.

- Solubility and stability of the test substance in the solvent/dispersant/vehicle/test medium:
The analysis of the test item/vehicle solutions of the 2.5%, 5% and 10% (w/w) concentrations for the actual test item levels was carried out (non-GLP) under conditions employing a validated analytical method. The analysis resulted in actual levels of 97.4 – 98.6% (2.5% (w/w) concentration), 96.1 – 98.5% (5% (w/w) concentration) and 97.4 – 100.2% (10% (w/w) concentration) of the nominal concentration.

Species:

mouse

Strain:

NMRI

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Sanhofer Weg 7, 97633 Sulzfeld, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 8-9 weeks
- Weight at study initiation: 26-34-g
- Housing:
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 12 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C  3°C (maximum range)
- Humidity (%): 55%  10% (maximum range)
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12 h/12 h
- IN-LIFE DATES: From: To: 07 November - 05 December 2019

Vehicle:

dimethyl sulphoxide

Concentration:

0, 2.5, 5, 10 %

No. of animals per dose:

6

Details on study design:

TREATMENT PREPARATION AND ADMINISTRATION:
The experimental schedule of the assay was as follows:
- Day 1:
The weight and any signs of toxicity of each animal were determined and recorded. In addition, ear swelling measurements were carried out at the helical edge of both ears using an Oditest micrometer.
Open application of 25 µL of the appropriate dilution of the test item, the vehicle alone or the positive control (as appropriate) were administered to the dorsum of each ear.
- Days 2 and 3:
The application procedure carried out on day 1 was repeated.
- Day 4 (24 hours after the last):
Ear swelling measurements (immediately before sacrificing the mice) were carried out at the helical edge of both ears using an Oditest micrometer.
The animals were euthanized by carbon dioxide (CO2) inhalation and laparotomised.
Punch biopsies of 8 mm in diameter of the apical area of both ears were prepared and immediately weighed on an analytical balance.
Lateral pairs of auricular lymph nodes draining the ear tissue were excised, carefully separated from remaining fatty tissue and weighed on an analytical balance immediately following preparation. The lymph nodes were then stored on ice in PBS /0.5% BSA and subjected to the preparation of single cell suspensions by mechanical tissue disaggregation. The cells were counted automatically in a cell counter.
Observations:
- weight of lymph nodes
- cell counts in lymph nodes
- stimulation index is calculated by dividing the absolute number of weight or cell counts of the substance treated lymph nodes by the vehicle treated ones
- ear swelling
- ear weight
- body weights
- clinical signs

PRE-SCREEN TESTS:
A preliminary experiment was carried out in 3 animals to examine the irritating potential and handling/application of the test item in order to select the appropriate concentrations. Three concentrations (w/w), 2.5%, 5% and 10% dissolved in DMSO were examined. Doses were selected according to OECD guideline from the concentration series 100%, 50%, 25%, 10%, 5%, 2.5%, 1%, 0.5% etc.
The preliminary experiment was conducted under conditions identical to the main LLNA study, except there was no assessment of lymph node proliferation and only 3 animals (one per concentration) were used.
- Irritation: No irritating properties were observed in this preliminary experiment at concentrations of 2.5%, 5% and 10%, no differences in ear weight and ear thickness were noted.
- Systemic toxicity: none
- Ear thickness measurements: yes

Positive control substance(s):

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

Statistics:

For lymph node weight a significance at p ≤ 0.01 is considered positive (U-test according to MANN and WHITNEY). A possible concentration-response-relationship was examined by linear regression analysis employing PEARSON's correlation coefficient. An U-test was performed also for the cell count.
In addition, the acute inflammatory skin reaction (irritating potential) was measured by ear weight determination of circular biopsies of the ears and ear thickness measurements on test day 1 and test day 4 to identify skin irritation properties of the test item employing the U-test according to MANN and WHITNEY by comparing the test groups to the vehicle control.

Positive control results:

Alpha hexyl cinnamic aldehyde (20% solution) shows a clear sensitizing potential in the local lymph node assay (IMDS).

Parameter:

SI

Remarks:

Stimulation Index

Value:

1

Test group / Remarks:

vehicle control group

Remarks on result:

other: no skin sensitization

Parameter:

SI

Remarks:

Stimulation index

Value:

0.996

Test group / Remarks:

2.5% test item

Remarks on result:

other: no skin sensitization

Parameter:

SI

Remarks:

Stimulation index

Value:

1.078

Test group / Remarks:

5% test item

Remarks on result:

other: no skin sensitization

Parameter:

SI

Remarks:

Stimulation index

Value:

0.985

Test group / Remarks:

10% test item

Remarks on result:

other: no skin sensitization

In the main study treatment with 1,12-Dodecanediol at the concentration of 2.5%, 5% and 10% test item did not reveal any statistical significantly increased values for the lymph node cell count and lymph node weight. The stimulation indices of the lymph node cell count did not exceed the threshold level of 1.4.

The threshold level for the ear weight of 1.1 was not exceeded and no increase of ear thickness was observed, i.e. no irritating properties were noted at any concentration.

The positive control group caused the expected increases in lymph node cell count and lymph node weight (statistically significant at p ≤ 0.01). Therefore, the study can be regarded as valid.

No signs of local or systemic intolerance were recorded.

parameter	control group (vehicle)	2.5% test item	5% test item	10% test item	positive control goup
lymph node cell count	1.000	0.996	1.078	0.985	1.602**
lymph node weight	1.000	1.036	1.164	1.291	1.519**
ear weight	1.000	0.939	0.862	0.981	1.065
ear thickness TD4	1.000	0.988	0.984	0.981	1.093

Interpretation of results:

other: no sensitizing potential; no indication for non-specific (irritant) activation

Executive summary:

The registered substance showed no sensitizing potential in the modified Local Lymph Node Assay (IMDS, following OECD TG 429) in female NMRI mice after dermal application of up to and including the maximum technically feasible concentration of 10 %. Additionally, no indication for a non-specific (irritant) activation was detected.

In conclusion, under the present test conditions, 1,12-Dodecanediol at the concentration of 2.5%, 5% or 10% (w/w) did not reveal any skin sensitising properties in the local lymph node assay and therefore 1,12-Dodecanediol is not classified to be skin sensitising in this test system.

Reference 2

Endpoint:

skin sensitisation: in vitro

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

August 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 442D (In Vitro Skin Sensitisation: ARE-Nrf2 Luciferase Test Method)

GLP compliance:

yes (incl. QA statement)

Type of study:

activation of keratinocytes

Specific details on test material used for the study:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The test item was completely dissolved in dimethyl sulfoxide (DMSO). Fresh preparations of the test were used for the treatment.

Details on the study design:

Skin sensitisation (In vitro test system) - Details on study design:
The study was conducted to investigate the potential of the test item to induce genes that are regulated by the antioxidant response element (ARE). The data may be used as part of an integrated approach to testing and assessment (IATA) to support the discrimination between skin sensitisers and non-sensitisers for the purpose of hazard classification and labelling.
The ARE-Nrf2 luciferase test method utilises an immortalised adherent cell line derived from HaCaT human keratinocytes. The cell line is stably transfected with a plasmid containing a luciferase gene under the transcriptional control of the SV40 promoter fused with the ARE from a gene known to be up-regulated by contact sensitisers.
The luciferase signal reflects the activation by sensitisers of endogenous Nrf2 dependent genes and the dependence of the luciferase signal in the recombinant cell line on Nrf2 has been demonstrated. This allows quantitative measurement (by luminescence detection) of luciferase gene induction, using well established light producing luciferase substrates, as an indicator of the activity of the Nrf2 transcription factor in cells following exposure to electrophilic substances.

Specifications:
KeratinoSens™ cell line supplied by Givaudan Schweiz, Zurich, Switzerland as specified in OECD Test Guideline 442D.

Preparation of Cultures:
For testing, cells were 80-90 % confluent, and care was taken to ensure that cells were never grown to full confluence. One day prior to testing cells were harvested and distributed into 96-well plates (10 000 cells/well). Attention was paid to avoid sedimentation of the cells during seeding to ensure homogeneous cell number distribution across wells. For each repetition, three technical replicates were used for the luciferase activity measurements, and three parallel technical replicates used for the cell viability assay.
A fresh vial of cells was used for each experimental occasion and cultured using Dulbecco’s modified Eagle medium (DMEM) containing 1 % serum and Geneticin.

Treatment:
In 96-well plates, incubated at 37±1 °C, 5 % (v/v) CO2, for about 48 hours in medium with serum but without Geneticin. For each test article and positive control, one experiment was needed to derive a prediction (positive or negative), consisting of at two independent repetitions each containing three replicates of each concentration. Each independent repetition was performed on a different day with fresh stock solution of test chemicals and independently harvested cells. Cells may come from the same passage however.

Luciferase Activity Measurements:
After the 48 hour exposure time with the test and control items, cells were washed with a phosphate buffered saline, and the relevant lysis buffer (One GlowTM Luciferase Assay System)6 for luminescence readings added to each well for an adequate time at room temperature. Plates with the cell lysate will then be placed in the luminometer for reading.

Cytotoxicity Assessment:
After the 48-hour exposure period, the medium was replaced with fresh medium containing MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and cells were incubated for 4 hours at 37 °C in the presence of 5 % CO2. The MTT medium was removed and cells were lysed by adding 10 % aqueous SDS solution to each well overnight or for up to 3 days at 37 °C. After shaking to ensure homogeneity of the solution in the wells and then absorption read at 620 nm using a photometer.

Positive controls:
Cinnamic aldehyde (CAS No. 14371-10-9), supplied by Sigma-Aldrich Chemie GmbH. was used as the positive control. A series of 5 master concentrations ranging from 0.4 to 6.4 mM was prepared in DMSO and diluted as described for the master concentrations, so that the final concentration of the positive control range from 4 to 64 μM.

Negative controls:
The negative control was diluted into culture medium containing serum so that the final concentration was 1 %. The solvent DMSO was used as the negative control, which is known to not affect cell viability and corresponds to the same concentration of DMSO found in the test item and in the positive control.

Positive control results:

The assay aceptance criteria for the positive controls were met:
Luciferase activity induction obtained with the positive control was statistically significant above the threshold of 1.5 at concentrations of 32 to 64 µM in Experiments 1 and 2.
The EC1.5 values for the positive control were 22.10 and 20.83 µM in Experiments 1 and 2, respectively. The average induction for the positive control at 64 µM was 2.33.

Run / experiment:

other: Exp. 1

Parameter:

other: Imax value

Value:

4.05

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Remarks on result:

positive indication of skin sensitisation

Run / experiment:

other: Exp. 2

Parameter:

other: Imax value

Value:

3.76

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Remarks on result:

positive indication of skin sensitisation

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: none
- Test item precipitation was noted macroscopically at concentrations of 125 µM and higher.

DEMONSTRATION OF TECHNICAL PROFICIENCY: proven

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes
- Acceptance criteria met for positive control: yes
- Acceptance criteria met for variability between replicate measurements: yes

The average coefficient of variation of the luminescence reading for the negative control (DMSO) was 6.72 % and 6.01 % in Experiments 1 and 2, respectively.

All assay acceptance criteria were met.

Since the Imax in both experiments was above 1.5-fold, the test article was considered to be positive in the ARE-Nrf2 Luciferase Test.

Table 1: Numerical results for the test item

	Luciferase determinations	Luciferase determinations	Cytotoxicity determinations	Cytotoxicity determinations
parameter	Imax	EC 1.5 [µM]	IC 50 [µM]	IC30 [µM]
test item, Exp. 1	4.05*	1.83*	430.50	363.58
test item, Exp. 2	3.76*	3.41*	397.89	319.37
average ± SD	3.91 ± 0.21	2.5  ± 1.12	413.87  ± 23.06	340.76  ± 31.26
positive control (64 µM) Exp. 1/2	2.44*/2.22*	22.10/20.83

* = significant compared to the negative control (p<0.05)

For the positive control cinnamic aldehyde the average fold induction in the two replicates at 64 μM should be between 2 and 8, the EC1.5 value should be between 7 μM and 30 μM.

Executive summary:

The study was conducted to investigate the potential of the test item to induce genes that are regulated by the antioxidant response element (ARE). The ARE-Nrf2 Luciferase Test Method (KeratinoSens) was performed acording to OECD 442D.

The test substance was solved in DMSO and tested at final concentrations of 0.98 to 2000 µM. The assay acceptance criteria for the positive control cinnamic aldehyde and the negative control DMSO were met in this test. Treatment of the cell cultures for 48 hours with the test item led to the following results: The maximum average fold induction of the luciferase activity (Imax) value observed at any concentration of the test item was 3.91 and the mean EC1.5 value, representing the concentration for which induction of luciferase activity is above the 1.5 fold threshold (i.e. 50 % enhanced luciferase acitivity), was 2.5 µM.

All assay acceptance criteria were met. Since the Imax values in both experiments were above 1.5-fold, the test article was considered to be positive in the ARE-Nrf2 Luciferase Test.

Reference 3

Endpoint:

skin sensitisation: in chemico

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

July 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 442C (In Chemico Skin Sensitisation: Direct Peptide Reactivity Assay (DPRA))

Version / remarks:

of June 2019

GLP compliance:

yes (incl. QA statement)

Type of study:

direct peptide reactivity assay (DPRA)

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL

- Solubility and stability of the test substance in the solvent/vehicle: analytically confirmed

FORM AS APPLIED IN THE TEST (if different from that of starting material): solution

Details on the study design:

Skin sensitisation (In chemico test system) - Details on study design:
The Direct Peptide Reactivity Assay (DPRA) is an in chemico procedure proposed to address the molecular initiating event leading to skin sensitization, namely protein reactivity, by quantifying the reactivity of test chemicals towards model synthetic peptides containing either lysine or cysteine. Cysteine and lysine percent peptide depletion values are then calculated and used in a prediction model to categorize a substance in one of four classes of reactivity for supporting the discrimination between skin sensitizers and non-sensitizers.
For comparison, tests were performed with the test item, the vehicle (solvent control = negative control) and the known sensitizer Cinnamic aldehyde (positive control).

The DPRA quantifies the remaining concentration of cysteine- or lysine-containing peptide following 24 hours incubation with the test item at 25 +/-2.5 ºC. Relative peptide concentration is measured by reversed phase (C18) high-performance liquid chromatography (HPLC) with gradient elution and UV detection at 220 and 258 nm. The synthetic peptides contain phenylalanine to aid in the detection.
The test item was dissolved and tested according to the given test procedure. Cinnamic aldehyde was used as positive control at a concentration of 100 mmol/L in acetonitrile.

Cysteine and lysine peptide solutions were incubated in glass autosampler vials with the test item at 1:10 and 1:50 ratio, respectively. The reaction solution was left in the dark at 25 ± 2.5°C for 24 ± 2 hours before running the HPLC analysis. The test item assay was analysed in triplicate for both peptides. Samples were visually inspected prior to HPLC analysis. The concentration of cysteine or lysine peptide was photometrically determined at 220 nm in each sample by measuring the peak area (area under the curve, AUC) of the appropriate peaks and by calculating the concentration of peptide using the linear calibration curve derived from the standards. Percent peptide depletion is calculated according to OECD Guideline.

The following criteria must be met for a run to be considered valid:
a) The standard calibration curve should have an r2 > 0.99.
b) The mean percent peptide depletion value of the three replicates for the positive control cinnamic aldehyde should be between 60.8% and 100% for the cysteine peptide and between 40.2% and 69.0% for the lysine peptide and the maximum standard deviation (SD) for the positive control replicates should be < 14.9% for the percent cysteine depletion and < 11.6% for the percent lysine depletion.
c) The mean peptide concentration of reference controls A should be 0.50 ± 0.05 mM and the coefficient of variation (CV) of peptide peak areas for the nine reference controls B and C in acetonitrile should be <15.0%.
If one or more of these criteria is not met, the run would have been repeated.

The following criteria must be met for a test item’s results to be considered valid:
a) The maximum standard deviation for the test item replicates should be < 14.9% for the percent cysteine depletion and < 11.6% for the percent lysine depletion.
b) The mean peptide concentration of the three reference controls C in the appropriate solvent should be 0.50 ± 0.05 mM.
If these criteria were not met, the data would have been rejected and the run have been repeated for that specific test item.

Positive control results:

Treatment with the positive control item revealed a cysteine and lysine peptide depletion of 69.83% for cysteine and 56.56% for lysine peptide. These values are within the required range of 60.8% and 100% for the cysteine peptide and between 40.2% and 69.0% for the lysine peptide. The maximum standard deviation (SD) for the positive control replicates were < 14.9% for the percent cysteine depletion and < 11.6% for the percent lysine depletion.

Run / experiment:

other: mean of 3 runs

Parameter:

other: % depletion in the cysteine 1:10/lysine 1:50 prediction model:

Value:

0.08

Vehicle controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: Reactivity Class: No or Minimal Reactivity DPRA prediction: negative

Run / experiment:

other: mean of 3 runs

Parameter:

other: % cysteine depletion

Value:

0.15

Vehicle controls validity:

valid

Positive controls validity:

valid

Parameter:

other: % lysine depletion

Value:

0

Vehicle controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

DEMONSTRATION OF TECHNICAL PROFICIENCY:
The acceptance criteria for a DPRA test to be considered valid were met.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for vehicle control: yes
- Acceptance criteria met for positive control: yes

The test item visually appeared a clear solution in acetone at the test concentration of 100 mmol/L.
No precipitate in the reaction mixture at the end of the incubation time and no co-elution were observed.

Results of the DPRA:

Test item	Mean % Cysteine peptide depletion	Mean % Lysine peptide depletion	Mean % Cysteine/Lysine peptide depletion	reactivity class	DPRA prediction
positive control (DNCB)	69.83	56.56	no data	moderate	positive
test item	0.15	0	0.08	no or minimal	negative*

Interpretation of results:

other: negative

Executive summary:

The Direct Peptide Reactivity Assay (DPRA; OECD 442C) is an in chemico procedure proposed to address the molecular initiating event leading to skin sensitization, namely protein reactivity, by quantifying the reactivity of test chemicals towards model synthetic peptides containing either lysine or cysteine. Cysteine and lysine percent peptide depletion values are then calculated and used in a prediction model to categorize a substance in one of four classes of reactivity for supporting the discrimination between skin sensitizers and non-sensitizers.

The test item was completely dissolved and stable in acetone at a concentration of 100 mM in the DPRA. No precipitate in the reaction mixture at the end of the incubation time and no co-elution were observed. The cysteine 1:10/lysine 1:50 prediction model was applied to the test item. No relevant depletion of cysteine and lysine peptides became obvious in the DPRA (0.08 %). According to the prediction model “no or minimal reactivity” was derived for the test item in acetone, leading to a DPRA prediction of “negative“.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Due to the physico-chemical characteristics of 1,12 -dodecanediol (i.e. solid, low water solubility: 18.7 mg/L, log Pow of 3.1 and a molecular mass of 202.33 g/mol) dermal uptake is not favoured (ECHA Guidance on Information Requirements, Chapter R7c, 2017).

1,12 -Dodecanediol was investigated in silico for structural activity relationship (QSAR), in chemico for peptide reactivity (DPRA), and in vitro for keratinocyte activation (ARE-Nrf2 Luciferase Test Method).

The structure activity relationship of the substance was investigated by using the OECD QSAR Toolbox 4.2.1 (released October 2018). No protein binding alerts for skin sensitization were identified in silico by the OECD QSAR Toolbox.

The Direct Peptide Reactivity Assay (DPRA; OECD 442C) is an in chemico procedure proposed to address the molecular initiating event leading to skin sensitization, namely protein reactivity, by quantifying the reactivity of test chemicals towards model synthetic peptides containing either lysine or cysteine. Cysteine and lysine percent peptide depletion values are then calculated and used in a prediction model to categorize a substance in one of four classes of reactivity for supporting the discrimination between skin sensitizers and non-sensitizers.

The test item was completely dissolved and stable in acetone at a concentration of 100 mM in the DPRA. No precipitate in the reaction mixture at the end of the incubation time and no co-elution were observed. The cysteine 1:10/lysine 1:50 prediction model was applied to the test item. No relevant depletion of cysteine and lysine peptides became obvious in the DPRA (0.08 %). According to the prediction model “no to minimal reactivity” was derived for the test item in water, leading to a DPRA prediction of “negative“.

The Luciferase Test Method (KeratinoSens) was performed acording to OECD 442D. The test substance was solved in DMSO and tested at final concentrations of 0.98 to 2000 µM. The assay acceptance criteria for the positive control cinnamic aldehyde and the negative control DMSO were met in this test. Treatment of the cell cultures for 48 hours with the test item led to the following results: The maximum average fold induction of the luciferase activity (Imax) value observed at any concentration of the test item was 3.91 and the mean EC1.5 value, representing the concentration for which induction of luciferase activity is above the 1.5 fold threshold (i.e. 50 % enhanced luciferase acitivity), was 2.5 µM. All assay acceptance criteria were met.

Since the Imax values in both experiments was above 1.5-fold, the test article was considered to be positive in the ARE-Nrf2 Luciferase Test.

The experimental investigation of the third key event of skin sensitization, the h-CLAT (OECD 442D) was initiated but terminated after the dose finding part of the assay. The test item appeared to be soluble in DMSO only at a reduced stock solution concentration of 100 mg/mL. However, after addition of the stock solution to the treatment medium the test item precipitated, could not be removed and disturbed the evaluation. Since the h-CLAT assay is not suitable for test items which precipitate into non stable suspensions or emulsions (defined by OECD 442E) the test item has to be considered as being outside the applicability domain of this test. The study could thus not be performed.

Due to the inconsistency of the results in silico (no alert), in chemico (negative) and in vitro KeratinoSens (positive) no final conclusion for skin sensitization could be drawn and the mouse LLNA was initiated.

The registered substance showed no sensitizing potential in the modified Local Lymph Node Assay (IMDS, following OECD TG 429) in female NMRI mice after dermal application of up to and including the maximum technically feasible concentration of 10 %. Additionally, no indication for a non-specific (irritant) activation was detected.

In conclusion, under the present test conditions, 1,12-Dodecanediol at the concentration of 2.5%, 5% or 10% (w/w) did not reveal any skin sensitising properties in the local lymph node assay and therefore 1,12-Dodecanediol should not be classified as skin sensitizer.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Due to inconsistency of the results for skin sensitization in chemico (negative) and in vitro (positive) no final conclusion on classification could be drawn. Thus, the mouse LLNA was initiated. The registered substance did not reveal any skin sensitising properties in the LLNA.

In conclusion, no classification for skin sensitization is warranted.