4-amino-2,5-dimethoxy-N-phenylbenzenesulphonamide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: well performed GLP and OECD guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

the relative humidity was between 30-75%. This did not affect the validity of the study

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

4-amino-2,5-dimethoxy-N-phenylbenzenesulphonamide

EC Number:

257-821-4

EC Name:

4-amino-2,5-dimethoxy-N-phenylbenzenesulphonamide

Cas Number:

52298-44-9

Molecular formula:

C14H16N2O4S

IUPAC Name:

4-amino-2,5-dimethoxy-N-phenylbenzene-1-sulfonamide

Details on test material:

- Name of test material (as cited in study report): Chinonbasesulfanilide TTR

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands B.V. Postbus 6174 NL - 5960 AD Horst / The Netherlands
- Age at study initiation: 7 - 12 weeks (beginning of acclimatisation)
- Weight at study initiation: average : 19.9 g
- Housing: single in Makrolon Type I, with wire mesh top
- Diet : pelleted standard diet, ad libitum
- Water : tap water, ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+-3
- Humidity (%): 30 - 75
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

6.25, 12.5, 25.0 % (in DMF)

No. of animals per dose:

4 per dose group

Details on study design:

Each test group of mice was treated by topical (epidermal) application to the dorsal surface of each ear lobe (left and right) with different test item concentrations of 6.25, 12.5, and 25 % (w/v) in DMF. The application volume, 25 µl, was spread over the entire dorsal surface (diameter: ca. 8 mm) of each ear lobe once daily for three consecutive days. A further group of mice was treated with an equivalent volume of the relevant vehicle alone (control animals).

3H-methyl thymidine (3HTdR) was purchased from GE Healthcare (specific activity, 2 Ci/mmol; concentration, 1 mCi/ml). Five days after the first topical application, all mice were administered with 250 µl of 81.18 µCi/ml 3HTdR (corresponds to 20.3 µCi 3HTdR per mouse) by intravenous injection via a tail vein.

Approximately five hours after treatment with 3HTdR all mice were euthanised by intraperitoneal injection of Na-thiopental.

The draining lymph nodes were rapidly excised and pooled per group (8 nodes per group). Single cell suspensions (in phosphate buffered saline) of pooled lymph node cells were prepared by gentle mechanical disaggregation through stainless steel gauze (200 µm mesh size). After washing two times with phosphate buffered saline (approx. 10 ml) the lymph node cells were resuspended in 5 % trichloroacetic acid (approx. 3 ml) and incubated at approximately +4 °C for at least 18 hours for precipitation of macromolecules. The precipitates were then resuspended in 5 % trichloroacetic acid (1 ml) and transferred to plastic scintillation vials with 10 ml of scintillation liquid and thoroughly mixed.

The level of 3HTdR incorporation was then measured on ß -scintillation counter. Similarly, background 3HTdR levels were also measured in two 1ml-aliquots of 5 % trichloroacetic acid. The ß-scintillation counter expresses 3HTdR incorporation as the number of radioactive disintegrations per minute (DPM).

Positive control substance(s):

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

Results and discussion

Positive control results:

Positive control substance: -Hexylcinnamaldehyde
Vehicle: acetone:olive oil (4+1)

Test Conc. / DPM / number of lymph nodes / DPM per ln / S.I.

0 % / 3088 / 8 / 386 / --
5% / 7489 / 8 / 936 / 2.43
10% / 12552 / 8 / 1569 / 4.07
25% / 11327 / 6 / 1883 / 4.88

EC3 = 6.7 %

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see compilation above

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Conc. (%) / DPM (background) / lymph nodes / DPM per lymph node / S.I control / 8293 / 8 / 1037 / -- 6.25 / 5982 / 8 / 748 / 0.72 12.5 / 5038 / 8 / 630 / 0.61 25.0 / 6657 / 8 / 824 / 0.80

Any other information on results incl. tables

No deaths occurred during the study period. No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period. The body weights of the animals, recorded prior to the 1st application and prior to necropsy was within the range commonly recorded for animals of this strain and age.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

The test item Chinonbasesulfanilide TTR was found to be not a skin sensitiser.

Executive summary:

In the study the test item Chinonbasesulfanilide TTR suspended in DMF was assessed for its possible contact allergenic potential.

For this purpose a local lymph node assay was performed using test item concentrations of 6.25, 12.5, and 25 %.

The animals did not show any clinical signs during the course of the study and no cases of mortality were observed.

In this study Stimulation Indices (S.I.) of 0.72, 0.61, and 0.80 were determined with the test item at concentrations of 6.25, 12.5, and 25 % in DMF, respectively.

The test item Chinonbasesulfanilide TTR was not a skin sensitiser in this assay. The EC3 value could not be calculated, since all S.I. were below 3.