1-Propanaminium, 2-hydroxy-N-(2-hydroxypropyl)-N,N-dimethyl-, esters with C18-unsatd. fatty acids, Me sulfates (salts)

Endpoint summary

Description of key information

Guinea pig maximisation test: Not sensitising (OECD guideline 406 and EU method B.6), Induction: intradermal; Challenge: topical)

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

Study period:

2011-01-17 to 2011-02-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Version / remarks:

(30 May 2008)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

(17 July 1992)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

A valid GPMT conducted according to guideline is available, which is reliable with restrictions and adequate for classification and labelling purposes. Potency estimation is not mandatory when existing guideline and GLP conforming data are available, which were conducted before the new annex of the REACH Regulation entered into force. Moreover, no indication for skin sensitisation was observed in this study, thus, no dose response information is needed. For this reason and for reasons of animal welfare no additional LLNA was conducted.

Specific details on test material used for the study:

- Name of test material: 1-Propanaminium, 2-hydroxy-N-(2-hydroxypropyl)-N,N-dimethyl-, esters with fatty acids, C18 unsatd., Me-sulfates (salts)
- Physical state: liquid
- Analytical purity: 100%

Species:

guinea pig

Strain:

other: Dunkin Hartley, HsdPoc:DH

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Age at study initiation: no data
- Weight at study initiation: 320.2 ± 14.0 g (test group) and 293.3 ± 21.2 g (control group)
- Housing: in groups of 2 or 3 animals in Makrolon type IV cages
- Diet (e.g. ad libitum): pelleted diet type "3023", Altromin International, Lage, Germany, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 13 days (for main experiment)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C ± 3°C
- Humidity (%): 30-70%
- Air changes (per hr): 8/h
- Photoperiod (hrs dark / hrs light): 12/12

Route:

intradermal and epicutaneous

Vehicle:

other: sesame oil

Concentration / amount:

Main experiment:
Intradermal: 0.5 % (w/w)
Dermal: 25 % (w/w)
Challenge: 1 % (w/w)

Route:

epicutaneous, occlusive

Vehicle:

other: sesame oil

Concentration / amount:

Main experiment:
Intradermal: 0.5 % (w/w)
Dermal: 25 % (w/w)
Challenge: 1 % (w/w)

No. of animals per dose:

10 (test group), 5 (control group)

Details on study design:

RANGE FINDING TESTS:
pilot experiment with 4 animals to determine which concentration of the test substance
- led to slight irritation after intradermal application (determination of the maximum compatible dose); concentrations tested: 5%, 3.5%, 2% and 0.5% (w/w)
- led to slight irritation after dermal application; concentrations tested: 100%, 75%, 50% and 25% (w/w)
- can just be applied dermally without leading to skin irritation (subirritative dose); concentrations tested: 10%, 5%, 1% and 0.5% (w/w)


MAIN STUDY
A. INDUCTION EXPOSURE
Intradermal induction (day 0)
- No. of exposures: 1
- Test groups: test substance
- Control group: vehicle
- Site: below the shoulder blades
- Frequency of applications: 1
- Concentrations:
0.1 mL FCA (mixed at a ratio of 1+1 in vehicle)
0.1 mL 0.5% test substance in sesame oil (or sesame oil in control)
0.1 mL 0.5% test substance (or sesame oil in control) + FCA (mixed at a ratio of 1+1 in sesame oil)

Topical induction (day 7)
- No. of exposures: 1
- Exposure period: 48 h
- Test groups: test substance
- Control group: sesame oil
- Site: below the shoulder blades, same region as for intradermal induction
- Frequency of applications: 1
- Concentrations: 25%

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 21
- Exposure period: 24 h
- Test groups: right flank: vehicle; left flank: test substance in Duhring chambers
- Control group: right flank: vehicle; left flank: test substance in Duhring chambers
- Site: flanks
- Concentrations: 1%
- Evaluation (hr after challenge): 24 h, 48 h

Challenge controls:

yes, vehicle only

Positive control substance(s):

yes

Remarks:

α-hexyl cinnamic aldehyde

Positive control results:

α-hexyl cinnamic aldehyde in sesame oil: sensitisation rate of 40% (4 of 10 animals positive); experiment from 2011-01-24 to 2011-02-18

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

1%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none reported

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none reported.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

1%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none reported

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none reported.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

1%

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none reported

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none reported.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

1%

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none reported

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 1%. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none reported.

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

α-hexyl cinnamic aldehyde in sesame oil

No. with + reactions:

4

Total no. in group:

10

Pilot experiment:

Animal	Application	Test substance concentration [%]	Scoring
1	Intradermal	5 3.5 2 0.5	3 3 2 1
2	Dermal without Duhring chambers	100 75 50 25	2 2 2 1
3	Dermal with Duhring chambers (challenge)	10 5	2 1
4	Dermal with Duhring chambers (challenge)	1 0.5	0 0

 

 

Main experiment:

Due to the intended activation of the immune system by FCA slight inflammation and subsequent crust formation was observed at the injection sites with FCA in the test group and the control group animals. This common response is not relevant for the grading.

After intradermal induction 10/10 animals of the test group showed discrete or patchy erythema (grade 1); no skin reactions were seen in the control group. After epicutaneous induction 5/10 animals of the test group showed discrete or patchy erythema (grade 1);no skin reactions were seen in 5/10 animals in the test group and in 5/5 animals in the control group.

After challenge no visible changes of the treated skin sites were observed in the test and control group animals 24 and 48 h after patch removal (= grade 0).

Interpretation of results:

not sensitising

Conclusions:

MDIPA Esterquat C18 unsatd. was not sensitising in this Guinea pig maximisation test.

Executive summary:

In a dermal sensitisation study according to OECD guideline 406 (17 July 1992) and EU method B.6 (30 May 2008) with MDIPA Esterquat C18 unsatd. (100% a.i.) 10 young adult Dunkin Hartley, HsdPoc:DH guinea pigs were tested using the method of method of Magnusson & Kligman (Guinea Pig Maximisation Test). The positive control α-hexyl cinnamic aldehyde produced a sensitisation rate of 40%.

Test concentrations were selected based on the results of a pilot study: intradermal induction 0.5% in sesame oil; epicutaneous induction 25% on sesame oil; challenge 1% in sesame oil. No pretreatment to create local skin irritation was necessary before epicutaneous induction as the selected concentration was slightly irritating.

After challenge no visible changes of the treated skin sites were observed in the test and control group animals 24 and 48 h after patch removal (= grade 0).

The test material produced a response in 0% of animals. According to CLP, EU GHS (Regulation (EC) No 1272/2008), a response of at least 30% of the test animals of an adjuvant type guinea pig test method for skin sensitisation is considered as positive.

MDIPA Esterquat C18 unsatd. is not a dermal sensitiser in this study.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Skin sensitisation

In a dermal sensitisation study according to OECD guideline 406 (17 July 1992) and EU method B.6 (30 May 2008) with MDIPA Esterquat C18 unsatd. (100% a.i.) 10 young adult Dunkin Hartley, HsdPoc:DH guinea pigs were tested using the method of method of Magnusson & Kligman (Guinea Pig Maximisation Test). The positive controlα-hexyl cinnamic aldehyde produced a sensitisation rate of 40%.

Test concentrations were selected based on the results of a pilot study: intradermal induction 0.5% in sesame oil; epicutaneous induction 25% on sesame oil; challenge 1% in sesame oil. No pretreatment to create local skin irritation was necessary before epicutaneous induction as the selected concentration was slightly irritating.

After challenge no visible changes of the treated skin sites were observed in the test and control group animals 24 and 48 h after patch removal (= grade 0).

The test material produced a response in 0% of animals. According to CLP, EU GHS (Regulation (EC) No 1272/2008), a response of at least 30% of the test animals of an adjuvant type guinea pig test method for skin sensitisation is considered as positive.

MDIPA Esterquat C18 unsatd. is not a dermal sensitiser in this study.

 

Also the source substances used for read-across MDEA-Esterquat C16-18 and C18 unsatd. and MDIPA-Esterquat C16-18 and C18 unsatd. were not sensitising. These supporting studies are included into the dossier to complete the toxicological profiles of teh substances an justify the read-across for other endpoints.

In dermal sensitisation studies according to OECD guideline 406 using the method of Bühler MDEA-Esterquat did not produce skin sensitisation. 

Similarly, in a dermal sensitisation study according to OECD Guideline 406 (GPMT) MDIPA-Esterquat C16-18 and C18 unsatd. was not a dermal sensitiser.

 

There is no data gap for skin sensitisation. Although no human data are available for the target substance MDIPA Esterquat C18 unsatd., there is no reason to believe that results obtained in guinea pigs would not be applicable to humans.

 

Respiratory sensitisation

MDIPA Esterquat C18 unsatd. did not show any skin sensitising properties in guinea pigs. There is no information available for respiratory sensitisation. Therefore, there is a data gap in this respect. However, the data gap cannot be fulfilled with experimental data, since there is no internationally accepted animal model for respiratory sensitisation. In case human data for respiratory sensitisation emerges, this will be taken into account.

 

Justification for the M&K test

The test substance MDIPA Esterquat C18 unsatd., is a surface active chemical with the alkyl moiety being predominantly composed of C18 unsaturated residues.

In recently published articles in peer reviewed journals (see reference list below, and publications cited in the revised OECD guideline 429 (July 2010)) it is clearly demonstrated that for the realistic assessment of the skin sensitization potential of surfactants the LLNA (OECD 429) is much less suitable than the Magnusson & Kligman method (OECD 406) and could lead to confounding results.

In particular, the OECD 429 guideline specifies that “In addition, test substance classes or substances containing functional groups shown to act as potential confounders (Basketter et al., 2009) may necessitate the use of guinea pig tests”.

Consequently, in the evaluation of such surface active substances for sensitizing properties, the LLNA test is not an appropriate assay and would not represent an optimum use of test animals. The mechanism underlying confounding or false positive result is not fully understood, but it may be that an unspecific, non-immunologically triggered mechanism may be the cause of an increased lymphocyte proliferation.

In contrast to the M&K, the LLNA focuses on the induction process of skin sensitization (i.e. lymphocyte proliferation), and does not capture the process of elicitation which checks if the organism had actually been sensitized or not. The design of the LLNA study does not allow to assess whether lymphocyte proliferation is a specific response or unspecific (false positive) response.

 

References:

Kreiling, R et al., Food and Chemical Toxicology 46 (2008): 1896-1904: Comparison of the skin sensitizing potential of unsaturated compounds as assessed by the local lymph node assay (LLNA) and the guinea pig maximization test (GPMT)

 

Basketter, D et al., Regulatory Toxicology and Pharmacology 55 (2009): 90-96: Application of a weight of evidence approach to assessing discordant sensitisation data sets: Implications for REACH

 

Garcia, C et al., Regulatory Toxicology and Pharmacology 58 (2010): 301-307: Comparative testing for the identification of skin sensitizing potentials of nonionic sugar lipid surfactants

 

Ball, N et al., Regulatory Toxicology and Pharmacology 60 (2011): 389-400: Evaluating the sensitizing potential of surfactants: integrating data from the local lymph node assay, guinea pig maximization test, and in vitro methods in a weight-of-evidence approach

 

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on relevant, reliable and adequate data MDIPA Esterquat C18 unsatd. has not to be classified and labelled according to the CLP Regulation (EC) No 1272/2008 with respect to skin sensitisation.