Reaction mass of: N,N'-[1,3-Phenylenebis(methylene)bis[12-hydroxyoctadecanamide], N,N'-hexane-1,6-diylbis(12-hydroxyoctadecanamide) and N,N'-ethane-1,2-diylbis(12-hydroxyoctadecan-1-amide)

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04. Apr. 2019 - 12. Jun. 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

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)

Test material

Test material information

Constituent 1

Reference substance name:

N,N'-ethane-1,2-diylbis(12-hydroxyoctadecan-1-amide)

EC Number:

204-613-6

EC Name:

N,N'-ethane-1,2-diylbis(12-hydroxyoctadecan-1-amide)

Cas Number:

123-26-2

Molecular formula:

C38H76N2O4

IUPAC Name:

12-hydroxy-N-[2-(12-hydroxyoctadecanamido)ethyl]octadecanamide

Test material form:

solid

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

Animal Health: The health condition of animals was examined by a veterinarian before initiation of the study. Animals were healthy, without visible signs of disease.
Acclimation: The animals were acclimated in identical conditions as during the experiment for 5 days prior to the start of treatment. The acclimation was according to standard operation procedures.
Housing Condition: The animals were housed in IVC polycarbonate cages (5 animals per cage) suspended on stainless steel racks, in a room equipped with central air-conditioning. The room temperature was within the range of 22 ± 3°C, relative humidity was within the range of 50 - 60 %. The light regimen was set to a 12-hour light / 12-hour dark cycle. The sanitation was performed according to standard operation procedures.
Diet: A laboratory food ssniff (ssniff Spezialdiäten GmbH, Germany) was served ad libitum, each day approximately at the same time.
Water: The animals received tap water for human consumption. Supply of drinking water was unlimited. The quality of drinking water is periodically monitored (including microbiological control) and recorded; certificate of analysis is included in raw data.
Bedding: Lignocel S3/4, Lufa - ITL GmbH, Germany
Animals Identification: Each animal was marked with permanent pen on the tail. Each cage was affixed with a cage card containing pertinent animal and study information.

Study design: in vivo (LLNA)

Vehicle:

dimethyl sulphoxide

Concentration:

100 % (w/v) (pre study)
25, 50, 100 % (w/v) (main study)

No. of animals per dose:

2 (pre study)
5 (main study)

Positive control substance(s):

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

Results and discussion

Positive control results:

SI: 5.57/valid and within the historical values of the testing facility

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

1.01

Test group / Remarks:

25 %

Results 2

Parameter:

SI

Value:

1.22

Test group / Remarks:

50 %

Results 3

Parameter:

SI

Value:

1.59

Test group / Remarks:

100 %

Results 4

Parameter:

EC3

Remarks on result:

not determinable

Remarks:

None of the SIs was higher than 3.

Cellular proliferation data / Observations:

In comparison with the control group, a moderate increasing of the pooled lymph node weights was observed at concentration of 100 %. The pooled lymph node weights of treated groups were 0.0350g for 25 % concentration, 0.0355 g for 50 % concentration and 0.0407 g for 100 % concentration of test item. The lymph node weight of control group and positive control group were 0.0370 g and 0.0734 g, respectively. The DPM values for the three treated groups were 1520.44 (25 %), 1830.86 (50 %) and 2393.11 (100 %), respectively. The SI values for the three treated groups were 1.01 (25 %), 1.22 (50 %) and 1.59 (100 %), respectively. The daily clinical observation of the animals did not show visible clinical signs of toxicity. Administration of the test item in used concentrations did not affect the body weight of treated animals. No erythema was observed in both mice in the pre test after test item administration.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

All validity criteria are fulfilled. The study can be considered as valid. The skin sensitizing potential of Bisamid was assessed using the murine local lymph node assay. Based on the results of this study, Bisamid is not considered a skin sensitizer under the condition of this LLNA study.

Executive summary:

The sensitization potential of Bisamid was evaluated using the Local Lymph Node Assay (LLNA). The LLNA has been developed to determine the allergic contact sensitization potential of chemicals. Based on the recommendations of the OECD Guideline 429, the test item was suspended in DMSO (w/v). The positive control (a-Hexylcinnamic aldehyde) (25 %) was dissolved in Acetone/Olive Oil 4:1. The Pre-screen testwas performed using the dose of 100 %. Based on the observations recorded in the Pre-screen tests, the concentration of 100 % was selected as top dose for the main test. Five female mice (CBA/Ca) per group were topically exposed (dorsum of both ears) to the test item at concentrations of 25 %, 50 % and 100 %, to the positive control and to the vehicle only. Lymphocyte proliferation was measured using incorporation of radioactive¹²⁵I-iododeoxyuridine and 10^-5M fluorodeoxyuridine in the draining lymph nodes. The radioactive incorporation was expressed as disintegrations per minute (DPM)/pooled treatment group and compared with DPM value from the vehicle control group and expressed as the Stimulation Index (SI). After application of the test itemat three concentrations (25 %, 50 % and 100 % w/v) the animals did not show visible clinical symptoms of either local irritation or systemic toxicity. In this study the Stimulation Indices (SI) of 1.01, 1.22 and 1.59 were determined with the test item at concentrations of 25 %, 50 %, and 100 % in DMSO, respectively. The EC3 value could not be calculated, since none of the tested concentrations induced a SI greater than the threshold value of 3. The test item Bisamid is not considered a skin sensitizer under the test conditions of this study.