Sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2000-10-23 to 2001-02-19

Reliability:

1 (reliable without restriction)

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

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)

Version / remarks:

1998

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate

EC Number:

403-080-9

EC Name:

Sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate

Cas Number:

92484-48-5

Molecular formula:

C16 H17 N3 O4 S .Na

IUPAC Name:

sodium 3-(2H-1,2,3-benzotriazol-2-yl)-5-(butan-2-yl)-4-hydroxybenzene-1-sulfonate

Details on test material:

- Name of test material: sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate (FAT 75'309/B)
- Physical state: Fine powder
- Analytical purity: >= 98%

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/Ca/Ola/Hsd

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Interfauna UK Limited, Blackthorne, Bicester, Oxon, UK
- Age at study initiation: Young adults
- Housing: A maximum of 4 mice was housed per cage, in cages suitable for animals of this strain and weight range.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: The animals were housed under the experimental conditions for at least 5 days, prior to the start of dosing.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3 degree C
- Humidity (%): 30 - 70 %
- Air changes (per hr): A minimum of 15 changes per hour
- Photoperiod: Artificial, giving 12 hours light, 12 hours dark

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Concentration:

25 µL of a 3%, 10% or 30% w/v preparation of the test substance in propylene glycol

No. of animals per dose:

4

Details on study design:

TREATMENT PREPARATION AND ADMINISTRATION:
Groups of four male mice were used for this study. Approximately 25µL of a 3%, 10% or 30% w/v preparation of the test substance in propylene glycol was applied, using a variable volume micro-pipette, to the dorsal surface of each ear. A vehicle control group was similarly treated using propylene glycol alone. The procedure was repeated daily for 3 consecutive days. Three days after the third application, all the animals were injected, via the tail vein, with approximately 250µL of phosphate buffered saline (PBS) containing approximately 20 µCi of a 2.0 Ci/mmol specific activity 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 degree C, the samples were pelleted by centrifugation and the supernatant was discarded. The cells were then resuspended in approximately 1 mL of TCA. The lymph node suspensions were transferred to scintillation vials and 10 mL of scintillant (Optiphase) was added prior to β-scintillation counting using a Packard Tri-Carb 2500TR Liquid Scintillation Counter.

Positive control substance(s):

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

Statistics:

Not performed

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

The application of the test substance at concentrations of 3%, 10% and 30% w/v in propylene glycol resulted in an isotope incorporation which was less than 3-fold at all three concentrations. The SI values were as follows: - 0.78 at a concentration of 3% test substance - 0.57 at 10% test substance - 2.06 at 30 % test substance

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: The disintegrations counted per minute were 1248 (3%), 915(10%) and 3283(30%) as compared to 1591 in the vehicle control group. Results for each group were obtained from 8 pooled auricular lymph nodes and show no dose response relationship.

Any other information on results incl. tables

Positive control

The application of hexylcinnamaldehyde at concentrations of 1%, 3% and 10% w/v in acetone resulted in a greater than 3-fold increase in isotope incorporation at all three concentrations. Therefore, hexylcinnamaldehyde was shown to be a skin sensitiser, confirming the validity of the protocol used for the study.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

In conclusion, the test substance is not considered to be a potential skin sensitizer.

Executive summary:

A local lymph node skin sensitisation test with the test substance sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate in mice was performed according to OECD Guideline 429. The assay determines the level of T lymphocyte proliferation in the lymph nodes draining the site of chemical application, by measuring the amount of radiolabelled thymidine incorporated into the dividing cells. The test substance was applied as 3%, 10% or 30% w/v preparations in propylene glycol. The application of the test substance at concentrations of 3%, 10% and 30% w/v in propylene glycol resulted in an isotope incorporation which was less than 3-fold at all three concentrations. Consequently, the test substance is not considered to be a potential skin sensitizer. In a positive control study, hexylcinnamaldehyde was shown to have the capacity to cause skin sensitisation when applied as 1%, 3% or 10% w/v preparations in acetone, confirming the validity of the protocol used for this study.