Amides, C12-18, N-[3-(dimethylamino)propyl]

Endpoint summary

Description of key information

Coco amidopropyldimethylamine was found not to lead to skin sensitisation when tested in guinea pigs according to the Buehler test method (OECD 406).

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-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 406 (Skin Sensitisation)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

Buehler test

Justification for non-LLNA method:

In recently published articles in peer reviewed journals, see reference list, it is clearly demonstrated that irritants and surfactants are more likely to give rise to false positives in the LLNA.
Consequently, in the evaluation of such substances for sensitizing properties the LLNA test is not an appropriate assay and would not represent an optimum use of test animals. It is therefore recommended that the Guinea Pig is used instead. This is also supported by the TG OECD 406 "In addition, test substance classes or substances containing functional groups shown to act as potential con founders (Basketter et al., 2009) may necessitate the use of guinea pig tests".

References:
1. Kreiling R et al. (2008).
Food Chem. Toxicol. 46. 1896-1904.
2. David Basketter, et al. (2009)
Regul Toxicol Pharmacol. 55: 90–96
3. Garcia C, et al. (2010)
Regul Toxicol Pharmacol. 58(2):301-307.
4. Nicholas Ball, et al. (2011)
Regul Toxicol Pharmacol. 60: 389–400

Species:

guinea pig

Strain:

Hartley

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories B.V. Kreuzelweg 53, 5961 NM/The Netherlands
- Age at receipt: 4 to 6 weeks old
- Weight at study initiation: 369-444 g (main study) 292-398 g (irritation screens)
- Housing: in groups of up to 10 animals during acclimatization, then individually housed in stainless steel cages
- Diet (e.g. ad libitum): ad libitum.Teklad Global Guinea pig diet 2040 C (batch No. 24/10) provided by Provimi Kliba AG as well as a haystick 4642 for environmental enrichment.
- Water (e.g. ad libitum): community tap water from Füllinsdorf ad libitum
- Acclimation period: 3 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3°C
- Humidity (%): 30-70%
- Air changes (per hr): 10-15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To: 2010-04-11 to 2010-24-12

Route:

epicutaneous, occlusive

Vehicle:

corn oil

Remarks:

provided by Carl Roth GmbH & CO.(Batch number 499150899)

Concentration / amount:

The test article was utilized as follows :

Irritation screen I :
at 100% and at 25%, 50% and 75% w/w in corn oil

Irritation screen II :
At 5% and at 0.1, 0.5 and 1% w/w in corn oil

Irritation screen III :
At 0.1% and at 0.01, 0.02 and 0.05% w/w in corn oil

Sensitization study
At 0.5% (induction) and 0.05% w/w and 0.01% w/w in corn oil (challenge).

Route:

epicutaneous, occlusive

Vehicle:

corn oil

Remarks:

provided by Carl Roth GmbH & CO.(Batch number 499150899)

Concentration / amount:

The test article was utilized as follows :

Irritation screen I :
at 100% and at 25%, 50% and 75% w/w in corn oil

Irritation screen II :
At 5% and at 0.1, 0.5 and 1% w/w in corn oil

Irritation screen III :
At 0.1% and at 0.01, 0.02 and 0.05% w/w in corn oil

Sensitization study
At 0.5% (induction) and 0.05% w/w and 0.01% w/w in corn oil (challenge).

No. of animals per dose:

Challenge : 20 test animals and 10 control animals
Irritation screens : 9 animals

Details on study design:

Treatment methods
The animal's fur was shaved with electric scissors just prior to the exposure. Closed patches were applied to the animals as follows:
0.5 mL of the undiluted test item or freshly prepared test item solution in a 25 mm Hill Top Chamber. Identical patching method was used for the irritation screens, induction and challenge(s). The Hill Top Chamber was firmly secured by an elastic plaster wrapped around the trunk of the
animal and secured with impervious adhesive tape. The occlusive dressing was left in place for six hours (± 15 minutes).

Irritation screens
For each irritation screen, 3 guinea pigs naïve animals were treated for approximately 6 hours.

Induction/ Performed on test Days 1, 8 and 15
The fur was clipped from the left flank of each test animal and the patches applied, over a period of 3 weeks. Each animal received one patch per week with the test item at 0.5% in corn oil, which remained in place for approximately 6 hours. The repeated application was performed at the same site. The interval between exposures was one week. The control animals remained untreated. After the last induction exposure the test animals were left untreated for 2 weeks before the challenge. Any gross skin reactions were recorded without depilation.

Challenge/ Performed on test Day 29
Test and control animals were challenged two weeks after the last induction with the test item at 0.05% and 0.01% in corn oil. The fur was clipped from the left posterior quadrant of the side and back of the animals. Patch sites for challenge are indicated below. The application period was 6 hours on a naive skin site.

Determination of Skin Reactions
The application area was depilated approximately 21 hours after the patches had been removed
(except during induction), using an approved depilatory cream (VEET Cream, Reckitt & Colman
AG, 4123 Allschwil / Switzerland). The depilatory cream was placed on the patch sites and
surrounding areas, and left on for up to 3-5 minutes. It was then thoroughly washed off with a
stream of warm, running water. The animals were then dried with a disposable towel, and
returned to their cages.
The grading method used for irritation screens, induction and challenge was identical. It was
performed 24 ± 2 hours after removal of the patches for irritation screens, induction and
challenge and repeated 24 ± 2 hours later (48-hour grades) for the irritation screens and the
challenge.
The scoring system was performed by visual assessment of erythema, oedema and other clinical
changes in skin conditions. They were assessed as follows:
0 = no visible change
1 = discrete or patchy erythema
2 = moderate and confluent erythema
3 = intense erythema and swelling
Grading of all animals was done by positioning each animal under true-light (Philips Master TLS
HE 28 W/840).
Note: At challenge, control animals were graded before the test animals.
For evaluation, two parameters were used: the incidence index and the severity index. The
incidence index is an expression of the number of animals showing a response of grade 1 or
greater at either 24- or 48-hour reading out of the total animals in the group, while the severity
index is calculated from the total sum of 24- and 48 hour responses divided by the number of
animals exposed.

Statistical Analysis
Descriptive statistics (means and standard deviations) were calculated for body weights.No
inferential statistics were used.

Challenge controls:

No treatment during the induction
Challenge animals
Test site number: 2
Concentration: 0.05 and 0.01% w/v in corn oil oil
Amount applied: 0.5 ml
Number of animals : 10

Positive control substance(s):

yes

Remarks:

alpha-hexylcinnamaldehyde (historical control)

Positive control results:

The sensitivity and reliability of the experimental technique employed was assessed by use of alpha-hexylcinnamaldehyde, which is recommended by the OECD 406 Guidelines and is known to have moderate skin sensitization properties in the guinea pig. The results from the most recent positive control test (Harlan Laboratories Study D06002, performed from 14-Sep-2010 to 10-Nov-2010) confirm alpha-hexylcinnamaldehyde as skin sensitizer, as it produced allergic contact dermatitis in >15% of the test animals.

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

0.5%

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 0.5%. No with. + reactions: 0.0. Total no. in groups: 20.0.

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. No with. + reactions: 0.0. Total no. in groups: 10.0.

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

0.5%

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 0.5%. No with. + reactions: 0.0. Total no. in groups: 20.0.

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. No with. + reactions: 0.0. Total no. in groups: 10.0.

Key result

Reading:

other: Conclusion

Hours after challenge:

48

Group:

positive control

Dose level:

No data

Remarks on result:

positive indication of skin sensitisation

Remarks:

The sensitivity and reliability of the experimental technique employed was assessed by use of alpha-hexylcinnamaldehyde, which is recommended by the OECD 406 Guidelines and is known to have moderate skin sensitization properties in the guinea pig. The results from the most recent positive control test (Harlan Laboratories Study D06002, performed from 14-Sep-2010 to 10-Nov-2010) confirm alpha-hexylcinnamaldehyde as skin sensitizer, as it produced allergic contact dermatitis in >15% of the test animals.

Range-finding study results

Due to the severe local signs, the animals used in the irritation screen I were euthanized for ethical reasons and a second irritation screen was performed with lower test item concentrations.

An induction concentration of 0.5% was selected for the induction because this level induced

only mild to moderate irritation in the second irritation screen. However, a non-irritating concentration could not be determined and therefore a third irritation screen was performed with lower test item concentrations.

Again, no non-irritating concentration could be determined in the irritation screen test. However, the results at all test item concentrations tested in the third irritation screen were similar. Therefore, the skin reactions were attributed to the vehicle (according to the experience of Harlan Laboratories Ltd., corn oil may have an irritating potential in guinea pig skin). Based on these results, a range of two test item concentrations was selected (0.05% and 0.01%) for the challenge to potentially achieve a fine distinction between skin irritation and sensitization.

Main study

Skin Reactions during Induction

Discrete or patchy erythema was observed in all test animals after treatment with the test item at 0.5% in corn oil.

 

Skin Reactions after Challenge

No skin reactions were observed in the control or test animals treated with the test item at 0.05% or 0.01% in corn oil.

 

Clinical Signs

The animals from the first irritation screen (nos. 1-3) were observed with swaying gait and

labored breathing after treatment. Otherwise, no clinical signs were observed during the course

of the study.

 

Body Weights

The body weight of the animals was within the range commonly recorded for this strain and age.

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Based on the above mentioned findings and in accordance with Regulation (EC) No 1272/2008, the substance does not have to be classified as skin sensitizer.

Executive summary:

The purpose of this skin sensitizing study was to assess the possible allergenic potential of cocoamidopropyldimethylamine when administered topically to albino Dunkin Hartley guinea pigs.

For this purpose the “Buehler Test” modified by Ritz, H.L. and Buehler, E.V. (1980) was used.Twenty male animals of the test group were treated topically with ARMAK 2096 at 0.5% in corn oil once a week for a 3-week induction phase. Two weeks after the final induction the animals were challenged with the test item at 0.05 and 0.01%.

The ten animals of the control group were not treated during the induction.They were challenged with the test item at 0.05 and 0.01%, identical to the test group animals.

 

None of the test or control animals were observed with skin reactions after the challenge with both test item concentrations. No clinical signs were recorded in the test or control animals during the course of the study. The body weights were within the range commonly recorded for this strain and age.

 

The sensitivity and reliability of the experimental technique employed was assessed by use of alpha-hexylcinnamaldehyde, known to have moderate skin sensitization properties in the guinea pig. The results from the most recent positive control test confirm alpha-hexylcinnamaldehyde as skin sensitizer, as it produced allergic contact dermatitis in >15% of the test animals.

 

Based on the above mentioned findings and in accordance with Regulation (EC) No 1272/2008, cocoamidopropyldimethylamine does not have to be classified as skin sensitizer.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

One skin sensitisation study is available. This skin sensitizing study was performed to assess the possible allergenic potential of coco amidopropyldimethylamine when administered topically to albino Dunkin Hartley guinea pigs. For this purpose the “Buehler Test” modified by Ritz, H.L. and Buehler, E.V. (1980) was used. Twenty male animals of the test group were treated topically with ARMAK 2096 at 0.5% in corn oil once a week for a 3-week induction phase. Two weeks after the final induction the animals were challenged with the test item at 0.05 and 0.01%. The ten animals of the control group were not treated during the induction.They were challenged with the test item at 0.05 and 0.01%, identical to the test group animals.

 

None of the test or control animals were observed with skin reactions after the challenge with both test item concentrations.No clinical signs were recorded in the test or control animals during the course of the study. The body weights were within the range commonly recorded for this strain and age.

 

The sensitivity and reliability of the experimental technique employed was assessed by use of alpha-hexylcinnamaldehyde, known to have moderate skin sensitization properties in the guinea pig. The results from the most recent positive control test confirm alpha-hexylcinnamaldehyde as skin sensitizer, as it produced allergic contact dermatitis in >15% of the test animals.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information:

As chemical respiratory sensitisers also elicit positive results in predictive tests for contact sensitisation, a negative outcome for dermal sensitisation is also predictive for non respiratory sensitisation of the substance.

Additionally, the likelihood for exposure via inhalation and thus becoming sensitised to cocoamidopropyldimethylamine is very low as result to the exposure via inhalation route. The substance is a solid with a melting point of 23°C that is marketed or used in a non solid or non granular form and with a vapour pressure around 3.96 10-5 Pa at 25°C (EPI suite estimation). Also the use of this substance will not result in aerosols, particles or droplets of an inhalable size, so exposure to humans via the inhalatory route will be unlikely to occur.

Justification for classification or non-classification

Cocoamidopropyldimethylamine has shown to be non-sensitising in a adequately performed Guinea pig study following the Buehler test. Consequently, the substance does not need to be classified for sensitisation.

These negative results for skin sensitisation can also be regarded as indicating the lack of the potential to cause allergic sensitisation of the respiratory tract.