Reaction products of C18 (unsaturated) fatty acids and dimethyl sulfate and triethanolamine

Endpoint summary

Description of key information

not sensitizing; weight of evidence: OECD TG 406 (Buehler test / Guinea pig maximisation test); RL1-2; GLP

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Reference 1

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1997-03-20 to 1997-05-30

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

17th July 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Version / remarks:

29 th Dec 1992

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

A valid GPMT conducted according to guideline is available, which is reliable without 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.

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male/female

Route:

intradermal and epicutaneous

Vehicle:

water

Concentration / amount:

Induction:
interdermal: 1 % test substance
dermal: 20 % test substance

Day(s)/duration:

dermal epicutaneous induction 7 d afer intradermal; 48 h exposure

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

Challenge: 1 % test substance
re-challenge: 1 % test substance

Day(s)/duration:

day 21 / day 28; 24 h exposure

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

Total of 55 animals (28 males and 27 females)
15 – preliminary study (2 animals for intradermal; 13 for topical)
20 – main study treatment group (10 males, 10 females)
10 – main study control (5 males, 5 females)
10 – main study reserve control for 2nd challenge
Route of administration intradermal and topical at induction; topical at challenge

Details on study design:

PRELIMINARY STUDY
A. Topical application
Irritation assessment following 24h occlusive exposure to test material at 100, 80, 60, 40, 20, 10, 5, 3, 1 and 0.5%; skin assessment 24 and 48h after removal of test substance.
Result: 20% was highest concentration to cause mild to moderate skin irritation. This concentration was selected for topical induction. No alterations were observed with 1% and 0.5%; 1% was selected for challenge.

B. Intradermal
0.1 ml of test substance at 0, 0.25, 0.5, 1, 3 and 5 % applied intradermally, both in physiological saline and in an emulsion made up of Freund’s Complete Adjuvant and physiological saline. Assessments made 24, 48 and 72h and 6 and 8 days post-administration Result: 1% in both vehicles (physiological saline and FCA) selected for intradermal induction.

MAIN STUDY
A. INDUCTION EXPOSURE
Day 0: Treatment group:
− Injection 1: 1:1 mixture (v/v) FCA/physiological saline
− Injection 2: 1% Tetranyl CO/40 in physiological saline
− Injection 3: 1% Tetranyl CO/40 in a 1:1 mixture (v/v) FCA/physiological saline
Control group:
- Injection 1: 1:1 mixture (v/v) FCA/physiological saline
- Injection 2: physiological saline
- Injection 3: 50 % (v/v) formulation of physiological saline in a 1:1 mixture (v/v) FCA/physiological saline

Day 7: Treatment group:
Region, free of fur, was treated topically with a 2 x 4 cm patch of filter paper, fully loaded with a 20% solution of Tetranyl CO/40 in bi-distilled water. Patch was covered by an occlusice bandage and left in place for 48 hours.
Control group:
Only vehicle was applied.

After removal of patches, test aereas were washed with warm water.


B. CHALLENGE EXPOSURE
Day 21: Left flank of all animals, including controls, treated topically with 0.5 ml of 1% test substance for 24h under an occlusive patch. 0.5 ml of vehicle alone applied to right flank.
Day 28: 2nd challenge. Test substance applied at 1% in bidest water to right flank and vehicle to left flank

GRADING SYSTEM
Dermal reactions graded for erythema and edema by blind reading according to grading scale:
No reaction: 0
Mild redness: 1
Moderate erythema: 2
Severe erythema with or without edema: 3

Positive control substance(s):

yes

Remarks:

2-Mercaptobenzothiazole

Positive control results:

The evaluation of the cutaneous alterations recorded in the treatment group, 24 and 48 h after the end of the exposure to the administered substance revealed that 10 of the 20 animals in the treatment group showed a reaction attributable to a sensibilisation process. the data are comparable to those obtained by Magnusson and Kligman (1969), for the assay with the substance Mercaptobenzothiazole

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

1 % test substance

No. with + reactions:

3

Total no. in group:

19

Remarks on result:

positive indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

48

Group:

test chemical

Dose level:

1 % test substance

No. with + reactions:

3

Total no. in group:

19

Remarks on result:

positive indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

1 % test substance

No. with + reactions:

1

Total no. in group:

10

Reading:

1st reading

Hours after challenge:

48

Group:

negative control

Dose level:

1 % test substance

No. with + reactions:

0

Total no. in group:

10

Reading:

rechallenge

Hours after challenge:

24

Group:

test chemical

Dose level:

1 % test substance

No. with + reactions:

0

Total no. in group:

19

Remarks on result:

no indication of skin sensitisation

Reading:

rechallenge

Hours after challenge:

48

Group:

test chemical

Dose level:

1 % test substance

No. with + reactions:

0

Total no. in group:

19

Remarks on result:

no indication of skin sensitisation

Reading:

rechallenge

Hours after challenge:

24

Group:

negative control

Dose level:

1 % test substance

No. with + reactions:

0

Total no. in group:

10

Reading:

rechallenge

Hours after challenge:

48

Group:

negative control

Dose level:

1 % test substance

No. with + reactions:

0

Total no. in group:

10

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

Mercaptobenzothiazole

No. with + reactions:

10

Total no. in group:

20

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

Mercaptobenzothiazole

No. with + reactions:

10

Total no. in group:

20

Preliminary Study

Topical application:

Result: 20% was highest concentration to cause mild to moderate skin irritation. This concentration was selected for topical induction. No alterations were observed with 1% and 0.5%; 1% was selected for challenge.

Intradermal injections:

Result: 1% in both vehicles (physiological saline and FCA) selected for intradermal induction

Main Study

Mortality:

One male, belonging to the treatment group, was found dead during the induction phase. But the death of this animal did not influence the results of the assay as the minimum number of animals required for the treatment group in assays in which Freud´s Complete Adjuvant is used during the induction phase is 10.

Grades:

1st challenge:

− Grade 1 reaction in 3/19 (15%) animals at 24 and 48h

− reactions in 1/10 (10 %) animals of control group at 24 h reading no effects at 48 h reading

2nd challenge:

- No effects in test or control group

Interpretation of results:

GHS criteria not met

Conclusions:

The test substance oleic acid-based TEA-Esterquat can be classified as a non-sensitiser to the guinea pig under the conditions of this study.

Executive summary:

In a dermal sensitization study with oleic acid-based TEA-Esterquat in bidest. water, young adult male and female Dunkin-Hartley guinea pigs were tested using the method of Magnusson and Kligman according to OECD guideline 406. Positive control material was 2-Mercaptobenzothiazole.

 In this study, the test substance oleic acid-based TEA-Esterquat produced a sensitisation rate of 15 % (3/19) at the first challenge and a sensitisation rate of 0 (0/20) at re-challenge at a concentration of 1 % and is considered to be a non-sensitizer.

Reference 2

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar (eco)toxicological properties because
• they share structural similarities with common functional groups: One quaternised ethanolamine moiety, one to three, mainly two ester groups with a typical UVCB distribution with long-chain fatty acids of natural origin. The molecular structure is almost identical.
• they are manufactured from similar resp. identical precursors (triethanolamine, long-chain fatty acids, dimethyl sulphate) under similar conditions. Therefore common breakdown products via physical and biological processes, which result in structurally similar chemicals are evident
• A constant pattern in the changing of the potency of the properties across the TEA-Esterquats by chain-length and the grade of esterification is not observed, because the fatty acid chain-length distribution is too narrow and similar and the distribution of mono-, di-, and tri-esters is identical. Some variation caused by variation in C=C double bonds may occur and will be discussed at the relevant endpoint.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See general justification for read-across attached to chapter 13 of this IUCLID file.

3. ANALOGUE APPROACH JUSTIFICATION
See general justification for read-across attached to chapter 13 of this IUCLID file.

4. DATA MATRIX
See general justification for read-across attached to chapter 13 of this IUCLID file.

Reason / purpose for cross-reference:

read-across: supporting information

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

1-10% in several studies

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

1-10% in several studies

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Group:

negative control

Remarks on result:

no indication of skin sensitisation

Group:

positive control

Dose level:

no details available

Remarks on result:

positive indication of skin sensitisation

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Reliable relevant data are available for oleic acid-based TEA-Esterquat and the structurally related substance partially unsaturated TEA-Esterquat. A justification for read-across is attached to iuclid section 13.

 

Summary

Two Buehler tests with partially unsaturated TEA-Esterquat were negative, one was judged as ambiguous due to an inconsistent response pattern and in most cases rapid fading of responses at challenge and re-challenge in test and control animals. In one further Buehler test the source substance partially unsaturated TEA-Esterquat was identified as weak sensitiser. Furthermore, two Guinea Pig Maximisation Tests, one conducted with partially unsaturated TEA-Esterquat, the other with oleic acid-based TEA-Esterquat are available, which were both negative.

A weight of evidence approach is used for evaluation. There is no indication from the available specifications to link the weak positive findings in the Buehler studies to differences in the degree of saturation of the underlying alkyl chain in partially unsaturated TEA-Esterquat.

The interpretation of results for those studies which were evaluated to be positive for skin sensitisation were complicated through the occurrence of a partly inconsistent pattern and in most cases rapid fading of response. Evaluation of response was hampered due to irritation and the absence of a clean negative control. Considering the studies with clean negative controls, proper performed induction with 100% test substance or mild to moderate irritating concentrations, there was no (0%) evidence for a skin sensitisation potential.

 

Table16: Reliable sensitisation studies with different methods

Method	fully saturated   TEA-EQ	partially unsaturated TEA-EQ	oleic acid-based  TEA-EQ
Bühler	2 negative	2 negative 1 ambiguous         1 weak sensitizer
Maximisation (Magnussen and Kligman)		1 negative	1 negative
Human HRIPT	2 negative studies with a total of 208 volunteers

  

 

Studies in detail

Skin sensitisation

In a dermal sensitization study according to OECD Guideline 406, May, 12 1981 with partially unsaturated TEA-Esterquat (a.i. 90 %, in isopropanol 10 %) diluted in water young adult Pirbright white guinea pigs (10 male, 10 female) were tested using the MAXIMIZATION TEST method.

Intradermal induction was performed with a 5 % test substance concentration. After dermal induction with 25 % solution of test substance, slight to well defined erythema and oedema was observed. None of the animals of the test group or control group showed any positive skin reactions after challenge treatment with a non-irritating test article concentration of 10 % in water. The sensitization rate at 24 h and at 48 h was 0 %. The test item is considered to be a non-sensitizer in this study.  

 

In a dermal sensitization study according to OECD Guideline 406, July 17, 1992 with partially unsaturated TEA-Esterquat (a.i. 90%) in water young adult Pirbright white guinea pig (10 male, 10 female) were tested using the BUEHLER method.

The sensitization rate at 24 h and at 48 h was 0 %. In this study, the test substance partially unsaturated TEA-Esterquat is considered to be a non-sensitizer. 

 

In a dermal sensitization study according to OECD Guideline 406, July, 17, 1992 with partially unsaturated TEA-Esterquat (a.i. 90 % in approx. 10 % isopropanol) diluted in purified water 20 male young adult Himalayan spotted guinea pigs were tested using the Bühler test method.

The sensitization rate at 24 h and at 48 h was 0 %. In this study, partially unsaturated TEA-Esterquat is considered to be a non-sensitizer. 

 

In a dermal sensitization study according to OECD Guideline 406, July, 17, 1992 with partially unsaturated TEA-Esterquat (a.i. 77 % in isopropanol) diluted in water 20 female young adult Pirbright-Hartley guinea pigs were tested using the Bühler test method.

For Induction a mild to moderate irritating concentration of 12.5 % was used. Challenge was performed with a 3 % dilution on the left and right flank. Grade 1 reactions were observed in 25, 55 and 5 % of test animals on right/ and or left flank at the 24, 48 and 72 hour reading, respectively. In 30 % of negative control animals as well a grade 1 reaction was observed at the 48 hour reading on left and/or right flank and in 10 % on the right flank at 72 hours. At re-challenge on the right flank with the same concentration grade 1 reaction was observed in 25, 45 and 15 % of test animals at 24, 48 and 72 hours, respectively. 20 and 30 % of control animals showed grade 1 reactions at 24 and 48 hours. 72 hours after application no effects were observed.

Re-challenge with 1 % revealed no reactions in negative control animals and no reactions on right flank of test animals. Grade 1 response was seen in 3/20 animals at the 48 hours reading, on the left flank. No reactions were observed at the 24 and 72 hour readings in test animals.

Considering only reactions with a clean control group and values of right flank as recommended by the OECD guideline 406 the results of the re-challenge with 1 % are negative with a sensitisation rate of 0 %. However, positive reactions have been observed at re-challenge with 1 % on the left flank, but only at one time point and with grade 1 in 3 animals. A partly inconsistent pattern and in most cases rapid fading of response was observed at challenge and re-challenge with 3 % test substance concentration in test and control animals. In conclusion the study results were evaluated to be ambiguous.

 

In a dermal sensitization study according to OECD Guideline 406, July, 17, 1992 with partially unsaturated TEA-Esterquat (a.i. 90 %) diluted in purified water 20 female young adult Pirbright-Hartley guinea pigs were tested using the Bühler test method.

Dermal induction was performed at concentrations of 60, 50 and 25 %, due to severity of response the concentration has to be decreased.

At challenge with 10 % test substance on the left and right flank, responses of slight erythema (score 0+, added by the study authors) and weak erythema/edema (score 1) were observed. No response was observed in negative control animals. For evaluation a response was considered positive, when a reaction on both flanks was observed, irrespective of severity. The resulting response rates were 25, 55 and 15 % at the 24, 48 and 72 hour readings, respectively. A consistent response over a period of 2 readings in minimum and a response on the contra lateral flank was observed in 35 % (7/20) of test animals. Re-challenge was performed with a 3 % dilution of the test substance on the left and right flank. Only responses on one flank were observed in 3 animals, most of them with a 0+ score. No reactions were observed in control animals. Due to the failure of response on the contra lateral flank the re-challenge was considered to be negative.

In conclusion there is evidence for a weak sensitisation potential of partially unsaturated TEA-Esterquat in this study.

 

In a dermal sensitization study with oleic acid-based TEA-Esterquat in bidest. water, young adult male and female Dunkin-Hartley guinea pigs were tested using the method of Magnusson and Kligman according to OECD guideline 406. Positive control material was 2-Mercaptobenzothiazole.

 In this study, the test substance oleic acid-based TEA-Esterquat produced a sensitisation rate of 15 % (3/19) at the first challenge and a sensitisation rate of 0 (0/20) at re-challenge at a concentration of 1 % and is considered to be a non-sensitizer.

 

There was no evidence for a skin sensitisation potential from two Human Repeated Insult Patch Tests involving a total of 208 volunteers.

 

Conclusion:

A weight of evidence approach covering all three TEA-Esterquats is used for evaluation. It is considered suitable to include all three TEA-Esterquats, differing mainly in the degree of saturation of fatty acid moiety, in the weight of evidence approach.

There is no indication from available specifications, to link the weak positive findings in theBühler studies to differences in the degree of saturation of the underlying alkyl chain in the partially unsaturated TEA-Esterquats.Therefore the result of this weight of evidence approach is justified to be valid without restrictions for this whole TEA-Esterquat group.

According to “Guidance on information requirements and chemical safety assessment Chapter R.7.3.4.2” reliable and relevant human data should be used for hazard identification and are even preferable to animal data. Adjuvant-type tests are likely to be more accurate in predicting a probable skin sensitizing potential of a substance in humans than those methods not employing Freund’s Complete Adjuvant and are thus the preferred of guinea pig methods (Council Regulation EC No 440/2008).

Under REACH, the LLNA is the preferred method for new developed skin sensitisation studies. Existing data of good quality deriving from guinea pig tests will be acceptable and will, if providing clear results, preclude the need for further in-vivo testing (Guidance on information requirements and chemical safety assessment Chapter R.7.3.3.1). Therefore no LLNA has been conducted with partially unsaturated TEA-Esterquat.

Considering the reliable studies, one study of the Bühler type was evaluated as weak positive and one of them was considered ambiguous because an inconsistent pattern of response was observed as outlined below. Further 4 Bühler studies and 2 guinea pig maximization tests were negative for skin sensitization as defined by GHS.

The interpretation of results for those studies which were evaluated to be positive for skin sensitisation were complicated through the occurrence of a partly inconsistent pattern and in most cases rapid fading of response. Evaluation of response was hampered due to irritation and the absence of a clean negative control. The studies rated with Klimisch 3 were not included in the evaluation,because no clear conclusion with regard to classification and labelling could be drawn. Considering the studies with clean negative controls, proper performed induction with 100 % test substance or mild to moderate irritating concentrations, there was no (0%) evidence for a skin sensitisation potential of the TEA-esterquats.

None of the 2 studies involving exposures of a total of 208 human volunteers to TEA-Esterquats was considered to induce a skin sensitisation response in humans.

The ability of chemicals to penetrate the human skin is a pre-requisite to cause a skin sensitisation response. Due to the relatively high molecular weight and physico-chemical properties the dermal penetration of TEA-Esterquats can be considered to be very low as outlined in chapter 5.1 toxicokinetics.

In conclusion, there was no evidence that the investigated TEA-Esterquats induced and/or elicited skin sensitisation responses in humans or the more accurate predicting adjuvant-type method in animals. Equivocal results were observed in the Bühler studies.Taking all evidence from available human and animal data together and emphasising the results of human data and data from guinea pig maximization test, the weight of evidence suggests, that TEA-Esterquats do not represent a skin sensitisation hazard and therefore none of the registered TEA- Esterquats need to be classified for skin sensitisation according to GHS Regulation EC No 1272/2008.

 

Similar results were obtained with the source substance MDEA-Esterquat C16-18 and C18 unsatd., results of Human Repeat Insult Patch Tests and supporting data from two Bühler studies with guinea pigs indicate that the substance does not induce skin sensitisation in humans. These data are included into the dossier to demonstrate, that both substances have a similar toxicological profile.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information:

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 or in vitro model for respiratory sensitisation. In case human data for respiratory sensitisation emerges, this will be taken into account.

Justification for classification or non-classification

Based on the available data, the substance does not need to be classified for skin sensitisation according to regulation (EC) 1272/2008. Thus, no labelling is required.