Butanal, reaction products with aniline

Endpoint summary

Description of key information

A modified Local Lymph Node Assay (IMDS) was performed on 24 female NMRI mice of the strain HsdWinin:NMRI (6 animals/test item group and 6 control animals) to determine if there is any specific (sensitizing) or non-specific (irritant) stimulating potential of the test item Butanal, reaction products with aniline.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

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:

guideline study

Qualifier:

according to guideline

Guideline:

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

Principles of method if other than guideline:

The modified Local Lymph Node Assay (IMDS) was performed on 24 female NMRI mice of the strain HsdWinin:NMRI(6 animals/test item group and 6 control animals) to determine if there is any specific (sensitizing) or non-specific (irritant) stimulating potential of the test item Butanal, reaction products with aniline.

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

NMRI

Sex:

male

Vehicle:

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

Concentration:

Group 1 Vehicle (A/OO - acetone/olive oil (4:1))
Group 2 3% Butanal, reaction products with aniline (in A/OO)
Group 3 10% Butanal, reaction products with aniline (in A/OO)
Group 4 30% Butanal, reaction products with aniline (in A/OO)
Group 5 30% Alpha Hexyl Cinnamic Aldehyde (in A/OO)

No. of animals per dose:

6 animals/test item group and 6 control animals

Parameter:

SI

Remarks:

Cell count index

Value:

1

Test group / Remarks:

Group 1: Vehicle (A/00)

Remarks on result:

other:

Remarks:

A modified Local Lymph Node Assay (IMDS) was carried out in mice. The modifications refer to the measurement of cell proliferation by cell counting instead of radioactive labeling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune competent cells in the draining lymph nodes, as also recommend in the update of OECD TG 429.

Parameter:

SI

Remarks:

Cell count index

Value:

1.02

Test group / Remarks:

Group 2: 3 % Butanal, reaction products with aniline (in A/00)

Remarks on result:

other:

Remarks:

A modified Local Lymph Node Assay (IMDS) was carried out in mice. The modifications refer to the measurement of cell proliferation by cell counting instead of radioactive labeling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune competent cells in the draining lymph nodes, as also recommend in the update of OECD TG 429.

Parameter:

SI

Remarks:

Cell count index

Value:

1.25

Test group / Remarks:

Group 3: 10 % Butanal, reaction products with aniline (in A/00)

Remarks on result:

other:

Remarks:

A modified Local Lymph Node Assay (IMDS) was carried out in mice. The modifications refer to the measurement of cell proliferation by cell counting instead of radioactive labeling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune competent cells in the draining lymph nodes, as also recommend in the update of OECD TG 429.

Parameter:

SI

Remarks:

Cell count index

Value:

1.41

Test group / Remarks:

Group 4: 30% Butanal, reaction products with aniline (in A/00)

Remarks on result:

other:

Remarks:

A modified Local Lymph Node Assay (IMDS) was carried out in mice. The modifications refer to the measurement of cell proliferation by cell counting instead of radioactive labeling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune competent cells in the draining lymph nodes, as also recommend in the update of OECD TG 429.

Parameter:

SI

Remarks:

Cell count index

Value:

1.45

Test group / Remarks:

Group 5: 30 %Alpha Hexyl Cinnamic Aldehyde (in A/00) - positive control

Remarks on result:

other:

Remarks:

A modified Local Lymph Node Assay (IMDS) was carried out in mice. The modifications refer to the measurement of cell proliferation by cell counting instead of radioactive labeling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune competent cells in the draining lymph nodes, as also recommend in the update of OECD TG 429.

Compared to vehicle treated animals there was an increase regarding the cell counts in the high dose group, which is of no statistical significance. The "positive level" of index 1.4 for the cell counts has been exceeded in this dose group.

The “positive level” of ear swelling, which is 2x10E-2 mm increase, i.e. about 10 % of the control values, has not been reached or exceeded in any dose group.

Nevertheless a statistically significant increase of the ear weight has been detected after administration of the mid concentration of the test item. Although this effect is not corroborated by increase in ear swelling an irritant potential cannot be excluded.

Direct LLNA (NMRI mice, female, 6 animals/group)

Groups	Weight index (index of mean+/- SD in%)	Cell count index (index of mean+/- SD in%)
Gr. 1	1.00 +/- 35.01	1.00 +/- 52.31
Gr. 2	0.99 +/- 24.05	1.02 +/- 31.64
Gr. 3	1.13 +/- 32.94	1.25 +/- 24.05
Gr. 4	1.15 +/- 10.62	1.41 +/- 22.41
Gr. 5	1.37 +/- 17.96	1.45 +/- 9.41

Ear swelling (NMRI mice, female, 6 animals/group, in 0.01 mm)

Groups	day 1 (mean+/- SD in%)	day 4 (mean+/- SD in%)	Index day 4
Gr. 1	17.33 +/- 2.84	18.17 +/- 7.73	1.00
Gr. 2	17.17 +/- 2.27	18.58 +/- 6.67	1.02
Gr. 3	17.42 +/- 4.55	19.50 +/- 5.99	1.07
Gr. 4	17.33 +/- 2.84	19.67 +/- 10.25	1.08
Gr. 5	17.67 +/- 4.41	19.92 +/- 3.36 *	1.09

* = statistically significant increase (p <= 0.05)

Ear weight (NMRI mice, female, 6 animals/group, in mg per 8 mm diameter punch)

Groups	day 4 (mean+/- SD in%)	Index day 4
Gr. 1	12.88 +/- 9.04	1.00
Gr. 2	13.64 +/- 10.68	1.06
Gr. 3	14.95* +/- 6.05	1.16
Gr. 4	13.82 +/- 12.78	1.07
Gr. 5	13.90 +/- 8.12	1.08

* = statistically significant increase (p <= 0.05)

Interpretation of results:

Category 1B (indication of skin sensitising potential) based on GHS criteria

Executive summary:

A modified Local Lymph Node Assay (IMDS) was performed on 24 female NMRI mice of the strain HsdWin:NMRI (6 animals/test item group and 6 control animals) to determine if there is any specific (sensitizing) or non-specific (irritant) stimulating potential of the test item Butanal, reaction products with aniline.

A concurrent control of 6 animals treated with Alpha Hexyl Cinnamic Aldehyde was included.

The study was conducted according to OECD Guidelines No. 429 and No. 406,

EC Guideline 2004/73/EC (29th Adaptation of Guideline 67/548/EEC, B.42)/Health Effects Test Guideline and OPPTS 870.2600 (EPA) with the following test item concentrations:

Test item: 0 % (vehicle control), 3 %, 10 % and 30 %.

Positive control: 30 % Alpha Hexyl Cinnamic Aldehyde

The test item and the positive control were formulated in acetone/olive oil (4:1) (A/OO) to yield a solution.

Compared to vehicle treated animals there was an increase regarding the cell counts in the high dose group, which is of no statistical significance. The "positive level" of index 1.4 for the cell counts has been exceeded in this dose group.

The “positive level” of ear swelling, which is 2x10E-2 mm increase, i.e. about 10 % of the control values, has not been reached or exceeded in any dose group.

Nevertheless a statistically significant increase of the ear weight has been detected after administration of the mid concentration of the test item. Although this effect is not corroborated by increase in ear swelling an irritant potential cannot be excluded.

In conclusion, these results show that the test item Butanal, reaction products with aniline may have a weak sensitizing potential in mice after dermal application of a 30 % concentration with an EC value of 28.75 %.

Therefore, the concentration of 10 % turned out to be the NOEL for the parameters investigated in this study with respect to skin sensitization.

These results are verified by the comparison with the results of the positive control group (Alpha Hexyl Cinnamic Aldehyde).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

The results of the modified LLNA showed that the test item Butanal, reaction products with aniline may have a weak sensitizing potential in mice after dermal application of a 30 % concentration with an EC value of 28.75 %.

Justification for selection of skin sensitisation endpoint:
Key study is used

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

In the LLNA an EC value of 28.75 % was determined. According to CLP classification criteria (Regulation (EC) No 1272/2008) as Skin Sens.1 B is justified.