Amines, C12-14-branched alkyl, dodecylbenzenesulfonates (1:1)

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24 March - 17 November 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study conducted in accordacen with international guidelines and in accordance with GLP. All guideline validity criteria were met.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

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:

Amines, C12-14-branched alkyl, octylbenzenesulfonates (1:1)

Molecular formula:

Amines, C12-branched alkyl, octylbenzenesulfonates: C27H51NO3S Amines, C13-branched alkyl, octylbenzenesulfonates: C28H53NO3S Amines, C14-branched alkyl, octylbenzenesulfonates: C29H55NO3S

IUPAC Name:

Amines, C12-14-branched alkyl, octylbenzenesulfonates (1:1)

Constituent 2

Reference substance name:

Amines, C12-14-branched alkyl, nonylbenzenesulfonates (1:1)

Molecular formula:

Amines, C12-branched alkyl, nonylbenzenesulfonates: C28H53NO3S Amines, C13-branched alkyl, nonylbenzenesulfonates: C29H55NO3S Amines, C14-branched alkyl, nonylbenzenesulfonates: C30H57NO3S

IUPAC Name:

Amines, C12-14-branched alkyl, nonylbenzenesulfonates (1:1)

Constituent 3

Reference substance name:

Amines, C12-14-branched alkyl, decylbenzenesulfonates (1:1)

Molecular formula:

Amines, C12-branched alkyl, decylbenzenesulfonates: C29H55NO3S Amines, C13-branched alkyl, decylbenzenesulfonates: C30H57NO3S Amines, C14-branched alkyl, decylbenzenesulfonates: C31H59NO3S

IUPAC Name:

Amines, C12-14-branched alkyl, decylbenzenesulfonates (1:1)

Constituent 4

Reference substance name:

Amines, C12-C14-branched alkyl, benzenesulfonates C12-C15 (1:1)

Molecular formula:

Amines, C12-branched alkyl, benzenesulfonates C12 (1:1) C31H59NO3S Amines, C12-branched alkyl, benzenesulfonates C13 (1:1) C32H61NO3S Amines, C12-branched alkyl, benzenesulfonates C14 (1:1) C33H63NO3S Amines, C12-branched alkyl, benzenesulfonates C15 (1:1) C34H65NO3S Amines, C13-branched alkyl, benzenesulfonates C12 (1:1) C32H61NO3S Amines, C13-branched alkyl, benzenesulfonates C13 (1:1) C33H63NO3S Amines, C13-branched alkyl, benzenesulfonates C14 (1:1) C34H65NO3S Amines, C13-branched alkyl, benzenesulfonates C15 (1:1) C35H67NO3S Amines, C14-branched alkyl, benzenesulfonates C12 (1:1) C33H63NO3S Amines, C14-branched alkyl, benzenesulfonates C13 (1:1) C34H65NO3S Amines, C14-branched alkyl, benzenesulfonates C14 (1:1) C35H67NO3S Amines, C14-branched alkyl, benzenesulfonates C15 (1:1) C36H69NO3S

IUPAC Name:

Amines, C12-C14-branched alkyl, benzenesulfonates C12-C15 (1:1)

Constituent 5

Reference substance name:

Amines, C12-14-branched alkyl, hexadecylbenzenesulfonates (1:1)

Molecular formula:

Amines, C12-branched alkyl, hexadecylbenzenesulfonates (1:1): C35H67NO3S Amines, C13-branched alkyl, hexadecylbenzenesulfonates (1:1): C36H69NO3S Amines, C14-branched alkyl, hexadecylbenzenesulfonates (1:1): C37H71NO3S

IUPAC Name:

Amines, C12-14-branched alkyl, hexadecylbenzenesulfonates (1:1)

Constituent 6

Reference substance name:

Residual Amines, C12-14-tert-alkyl

IUPAC Name:

Residual Amines, C12-14-tert-alkyl

Test material form:

liquid: viscous

Details on test material:

Storage: Room Temperature

Specific details on test material used for the study:

RADIOLABELLING INFORMATION (if applicable)
- Radiochemical purity: N/A
- Specific activity: N/A
- Locations of the label: N/A
- Expiration date of radiochemical substance: N/A

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: 15-25 ºC, protected from light
- Stability under test conditions: Assumed stable
- Solubility and stability of the test substance in the solvent/vehicle: Soluble in acetone: olive oil 80:20 v/v
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: No

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Dissolved in acetone: olive oil 80:20 v/v
- Preliminary purification step (if any): N/A
- Final dilution of a dissolved solid, stock liquid or gel: Test item dose solutions prepared at 50, 25, 10 % in acetone: olive oil 80:20 v/v
- Final preparation of a solid: N/A

FORM AS APPLIED IN THE TEST (if different from that of starting material) : Applied as a liquid

TYPE OF BIOCIDE/PESTICIDE FORMULATION (if applicable) : N/A

OTHER SPECIFICS: N/A

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaCrl

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River, Margate, UK
- Females (if applicable) nulliparous and non-pregnant: Yes
- Microbiological status of animals, when known: Not reported
- Age at study initiation: Not reported
- Weight at study initiation: 18-23 g, individual weight body weights were within ±20 % of the mean
- Housing: The animals were housed in groups of up to five during acclimatisation in period in suspended, solid floor cages with wire lids. From Day-1, the preliminary study animal was individually housed and the main study animals were housed in groups of up to three. Bedding was provided on a weekly basis to each cage by use of clean European softwood bedding (Datesand Ltd., Manchester, UK). The bedding had been analysed for specific contaminants and the results retained on file at Covance. No contaminants were present in bedding at levels which might have interfered with achieving the objective of the study.
- Diet (e.g. ad libitum): 5LF2 EU Rodent Diet 14%, was freely available to the animals at all times. Each batch of diet had been analysed for specific constituents and contaminants by the manufacturer. No contaminants were present in diet at levels which might have interfered with achieving the objective of the study. Results are retained on file at Covance.
- Water (e.g. ad libitum): Mains water was provided, ad libitum, via cage-mounted water bottles. The water had been periodically analysed for specific contaminants. No contaminants were present in water at levels which might have interfered with achieving the objective of the study. Results are retained on file at Covance.
- Acclimation period: 22 days
- Indication of any skin lesions: No

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24 ºC
- Humidity (%): 45-65 %
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12:12 light:dark
- IN-LIFE DATES: From: 28 March 2017 To: 25 April 2017

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

10, 25, 50 % w/v

No. of animals per dose:

5

Details on study design:

PRE-SCREEN TESTS:
- Compound solubility: Soluble in vehicle at 50 % w/v
- Irritation: Very slight (barely perceptible) erythema was noted from Day 2 to Day 6 and there was no change in ear thickness.
- Systemic toxicity: No evidence of systemic evidence
- Ear thickness measurements: No change in ear thickness
- Erythema scores: 0 (Day 1) and 1 (Day 2 through 6)

MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Mouse Local Lymph Node Assay
- Criteria used to consider a positive response: The proliferation response of lymph node cells was expressed as the number of radioactive disintegrations per minute per animal and as the ratio of 3HTdR incorporation into lymph node cells of test nodes relative to that recorded for the control nodes (Stimulation Index).

The test item will be regarded as a sensitizer if at least one concentration of the test item results in a threefold or greater increase in 3HTdR incorporation compared to control values. Any test item failing to produce a threefold or greater increase in 3HTdR incorporation will be classified as a "non sensitizer".


TREATMENT PREPARATION AND ADMINISTRATION:

Formulations were freshly prepared as required using 80% v/v acetone in olive oil on Days 1, 2 and 3. The formulations were stored at room temperature in sealed, air- tight containers prior to dosing and were used within two hours of preparation. The formulations were mixed by multiple inversion of the containers prior to administration to ensure homogeneity.

Concentrations of test article were expressed gravimetrically and in terms of test article received (without regard to purity or active content).

The five groups of five female mice were subjected to application of the vehicle control, positive control or one of the test formulations to the outer aspect of the auditory pinnae, once daily on Days 1, 2 and 3.

Positive control substance(s):

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

Results and discussion

Positive control results:

The positive control article produced a Stimulation Index of 17.27 (positive indication)

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

17.22

Test group / Remarks:

10 % w/v

Results 2

Key result

Parameter:

SI

Value:

24

Test group / Remarks:

25 % w/v

Results 3

Key result

Parameter:

SI

Value:

25.16

Test group / Remarks:

50 % w/v

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA

Not reported

DETAILS ON STIMULATION INDEX CALCULATION

The scintillation counter provided data including the DPM value (disintegrations per minute during a ten minute period) for each individual animal. The mean DPM value for each test group was divided by the mean DPM for the control group to provide the Stimulation Index (SI) value for each test group.

EC3 CALCULATION

Not determined.

CLINICAL OBSERVATIONS:

There were no clinical signs indicative of a systemic effect of treatment among mice treated with the vehicle or with 10, 25 or 50 % w/v formulations of the test article. Very slight erythema was noted on the backs of the ears of all animals treated at 50 % w/v from Day 2 to Day 6 and in animals treated at 25 % w/v from Day 3 to Day 6. No irritation was noted in the vehicle control group, the positive control group or in animals treated at 10 % w/v. Greasy fur to the neck and back of ears was noted in all test groups and the positive control group from Day 2 to Day 6.

BODY WEIGHTS

There was no indication of a treatment related effect on body weight.

Any other information on results incl. tables

Table 1      Stimulation index results

 

Test Group	Animal #	Individual animal DPM	Mean group DPM (std. dev.)	Stimulation index (SI)^a
Solvent control (acetone: olive oil 4:1 v/v)	104	475	283 (130.8)	N/A
	105	107
	106	271
	107	265
	108	295
10 % test item	109	4926	4866 (1785.6)	17.22
	110	6888
	111	1979
	112	5236
	113	5301
25 % test item	114	7923	6783 (843.2)	24.00
	115	5721
	116	6807
	117	7184
	118	6278
50 % test item	119	6074	7112 (711.8)	25.16
	120	7955
	121	7516
	122	6844
	123	7169
Positive control	124	5672	4881 (1034.7)	17.27
	125	5500
	126	4320
	127	5597
	128	3317

^aStimulation Index (SI) of ≥ 3.0 indicates a positive result

Applicant's summary and conclusion

Interpretation of results:

Category 1 (skin sensitising) based on GHS criteria

Conclusions:

The test article was classified as a Category 1 sensitiser according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS).

Executive summary:

OECD 429 (2017) - In a dermal sensitization study with Amines, C12-14-branched alkyl, dodecylbenzenesulfonates (1:1) in acetone: olive oil (4:1 v/v) young female adult mice (CBA/CaCrl) were tested using the Local Lymph Node Assay (LLNA)

 

Following a preliminary screening test in which no clinical signs of toxicity were noted at a concentration of 50 %, this concentration was selected as the highest dose investigated in the main test. Three groups, each of five animals, were treated with 25 μL of the test item in solution in the vehicle at concentrations of 50, 25 and 10 % w/v. A further group of five animals was treated with the vehicle alone. A concurrent positive control test, using a group of five animals, was also performed with the known sensitiser, α-Hexylcinnamaldehyde at a concentration of 25 % v/v in the vehicle.

 

There were no signs of systemic toxicity through the test period. Local skin irritation observed was limited to grade 1 erythema i.e. very slight erythema (barely perceptible) and ear thickness were within normal range. There were no clinical abnormalities or macroscopic abnormalities of the surrounding area were noted for any of the animals and no mortality reported during the study. The Stimulation Index, expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group, was determined for each treatment group.

 

The test item did elicit a Stimulation Index ≥ 3 when tested at ≥ 10 % w/v with SI values of 17.22, 24.00 and 25.16 in the 10, 25 and 50 % w/v test groups, respectively. The test item was therefore considered to be a sensitiser under the conditions of the test.

 

In this study, Amines, C12-14-branched alkyl, dodecylbenzenesulfonates (1:1) was a dermal sensitiser.

 

Based on the condition of this study, the test item was classified as a Category 1 sensitiser according to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS).