Potassium Lauroyl Wheat Amino Acids

Endpoint summary

Description of key information

Based on the results of the read across studies, the test substance is considered as non-sensitising to the skin.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

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

Adequacy of study:

key study

Study period:

From May 07, 2001 to August 23, 2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

KL2 due to RA; A read-across study with an analogous substance was scientifically justified, because it was conducted from the disodium salt to the monosodium salt of the test substance.

Justification for type of information:

Refer to section 13 of IUCLID for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

Buehler test

Justification for non-LLNA method:

Once the LLNA became an OECD method, the Buehler was already performed. Furthermore, the substance is irritating and for irritating substances a LLNA can be waived.

Species:

guinea pig

Strain:

other: Himalayan spotted

Sex:

male

Details on test animals and environmental conditions:

Test animals
- Source: RCC Ltd, Biotechnology & Animal Breeding Division, Wolferstrasse 4, CH-4414 Fullinsdorf / Switzerland
- Age at study initiation: 4-6 weeks
- Weight at study initiation: 290-418 g
- Housing: Makrolon type-4 cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: one week

Environmental conditions
- Temperature (°C): 22 ±3°C
- Humidity (%): 30-70%
- Air changes (per h): 10-15
- Photoperiod (h dark / h light): 12h/12h

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

undiluted

Day(s)/duration:

Week 1, Week 2 and Week 3, 6 h exposure each week

Adequacy of induction:

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

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

other: bi-distilled water

Concentration / amount:

50%

Day(s)/duration:

2 weeks after last induction

Adequacy of challenge:

highest non-irritant concentration

No.:

#2

Route:

epicutaneous, occlusive

Vehicle:

other: bi-distilled water

Concentration / amount:

25 and 50% (Rechallenge test) purified test substance

Day(s)/duration:

2 weeks after challenge test

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

Test group: 20 animals
Control group: 10 animals

Details on study design:

Irritation screening test:
An irritation screening test was performed to determine the minimal irritating concentration used in the induction period and the highest non-irritating concentration used for the challenge. Four different concentrations (5%, 25%, 50%, 100% diluted with bi-distilled water) were used on each animal for a 6-h exposure period. 4 guinea pigs were used. Application sites were assessed after 24 and 48 h. The most representative concentration to stimulate a state of immune hypersensitivity was 100 % used in the induction phase and a concentration of 50% was used in the challenge as the highest non-irritating concentration.

Main study
1) Induction
The fur was clipped from the left shoulder of each test animal and the patches applied, over a period of 3 weeks. The animals were treated with the test substance applied undiluted. Each animal received one patch per week which remained in place for approximately 6 h each. The repeated application was performed at the same site. The interval between exposure 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. The skin responses were graded 24 h after the patches had been removed. Any gross skin reactions were recorded without depilation.

2) Challenge- performed on test Day 29
The animals previously exposed during the induction period (i.e. test group) as well as the previously untreated control animals were challenged two weeks after the last induction exposure using the test substance at 50 % in bi-distilled water. The fur was clipped from the left posterior quadrant of the side and back of the animals. The exposure period was 6 h on a naive skin site. The responses were graded at 24 and 48 h after the patches had been removed.

3) Second challenge
The test group was rechallenged 14 d following primary challenge. All animals in the test group were included in the rechallenge. The test substance was applied on the right cranial flank at 50 % in bi-distilled water and on the the right caudal flank at 25 % in bi-distilled water. The grading method used for irritation screen, induction and challenge was identical. 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 TLD 36W/84 or Osram 36W/31 830).

Challenge controls:

During induction the control animals remained antreated. At the challenge the controls were treated the same as the test group. Only the test group was challenged a second time.

Positive control substance(s):

yes

Remarks:

2-Mercaptobenzothiazole

Positive control results:

Five out of 10 test animals were observed with discrete/patchy erythema at the 24-h reading and moderate/confluent erythema was observed at the 48-h reading in all ten test animals after the challenge treatment with the highest tested non-irritating concentration of 2-Mercaptobenzothiazole at 0.03 % in mineral oil. No skin reactions were observed in the control group.

Key result

Reading:

rechallenge

Hours after challenge:

24

Group:

test chemical

Dose level:

25% test substance

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Key result

Reading:

rechallenge

Hours after challenge:

48

Group:

test chemical

Dose level:

25% test substance

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

bi-distilled water

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

bi-distilled water

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.03% 2-Mercaptobenzothiazole in mineral oil

No. with + reactions:

5

Total no. in group:

10

Clinical observations:

discrete/patchy erythema

Remarks on result:

positive indication of skin sensitisation

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.03% 2-Mercaptobenzothiazole in mineral oil

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

moderate/confluent erythema

Remarks on result:

positive indication of skin sensitisation

In the first challenge application 12 out of 20 animals reacted with discrete erythema after 24 and 48 h. the challenge was repeated with a purified sample in the sensitized animals. None of the animals reacted to the purified sample. The purification step is always applied to the commercial product.

Interpretation of results:

other: not classified based on EU CLP criteria

Conclusions:

Based on the results of the read across study, the test substance is considered to be non-sensitising to the skin.

Executive summary:

A study was conducted to determine skin sensitisation potential of read across substance, sodium cocoyl glutamate, using the Buehler method, according to OECD Guideline 406, in compliance with GLP. The test was performed in 20 (10 test and 10 control) male Himalayan spotted guinea pigs. Based on the screening test, 100% (undiluted) and 50% (in bi-distilled water) test substance concentrations were selected for induction and challenge phase, respectively. In the induction phase, fur was clipped from the left shoulder of each test animal and occlusive patches of the undiluted test substance applied (one a week at the same site) for 6 h over a period of 3 weeks. The skin responses were graded 24 h after the patches had been removed. Any gross skin reactions were recorded without depilation. The control animals remained untreated. After a rest period of 2 weeks, both the test and control group animals were challenged on a naive skin site with 50% test substance in bi-distilled water under similar conditions. The responses were graded at 24 and 48 h after the patches had been removed. All animals in the test group were additionally re-challenged on the right cranial flank at 50% in bi-distilled water and on the right caudal flank at 25% in bi-distilled water. The scoring system was performed by visual assessment of erythema, oedema and other clinical changes in skin conditions. They were assessed as: 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 TLD 36W/84 or Osram 36W/31 830). During challenge 12 out of 20 animals reacted with distinct erythema. Therefore, a purified sample was prepared (which is equivalent to the commercial form) for the re-challenge procedure. During re-challenge with the purified sample no skin reactions were observed after 24 and 48 h in any animal. In the positive control group, five out of ten test animals were observed with discrete/patchy erythema at the 24-h reading and moderate/confluent erythema was observed at the 48-h reading in all ten test animals after the challenge treatment with the highest tested non-irritating concentration of 2-Mercaptobenzothiazole at 0.03 % in mineral oil. No skin reactions were observed in the negative control group. Under the study conditions, the read across substance was determined to be non-sensitising to the skin (BASF, 2001). Based on the results of the read across study, similar absence of skin sensitisation potential can be expected for the test substance, 'potassium lauroyl wheat amino acids'.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information:

In absence of skin sensitisation study with the test substance, the endpoint has been assessed based on studies for substances represenative of the main constituents, which can be chemically categorised as ‘acyl amino acid salts or amino acid alkyl amides, together with expert opinions on hydrolysed wheat proteins in general. The results are presented below:

Studies on amino acid alkyl amides:

Study 1: A study was conducted to determine skin sensitisation potential of read across substance, 'sodium cocoyl glutamate', using the Buehler method, according to OECD Guideline 406, in compliance with GLP. The test was performed in 20 (10 test and 10 control) male Himalayan spotted guinea pigs. Based on the screening test, 100% (undiluted) and 50% (in bi-distilled water) test substance concentrations were selected for induction and challenge phase, respectively. In the induction phase, fur was clipped from the left shoulder of each test animal and occlusive patches of the undiluted test substance applied (one a week at the same site) for 6 h over a period of 3 weeks. The skin responses were graded 24 h after the patches had been removed. Any gross skin reactions were recorded without depilation. The control animals remained untreated. After a rest period of 2 weeks, both the test and control group animals were challenged on a naive skin site with 50% test substance in bi-distilled water under similar conditions. The responses were graded at 24 and 48 h after the patches had been removed. All animals in the test group were additionally re-challenged on the right cranial flank at 50% in bi-distilled water and on the right caudal flank at 25% in bi-distilled water. The scoring system was performed by visual assessment of erythema, oedema and other clinical changes in skin conditions. They were assessed as: 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 TLD 36W/84 or Osram 36W/31 830). During challenge 12 out of 20 animals reacted with distinct erythema. Therefore, a purified sample was prepared (which is equivalent to the commercial form) for the re-challenge procedure. During re-challenge with the purified sample no skin reactions were observed after 24 and 48 h in any animal. In the positive control group, five out of ten test animals were observed with discrete/patchy erythema at the 24-h reading and moderate/confluent erythema was observed at the 48-h reading in all ten test animals after the challenge treatment with the highest tested non-irritating concentration of 2-Mercaptobenzothiazole at 0.03 % in mineral oil. No skin reactions were observed in the negative control group. Under the study conditions, the read across substance was determined to be non-sensitising to the skin (BASF, 2001).Based on the results of the read across study, similar absence of skin sensitisation potential can be expected for the test substance, 'potassium lauroyl wheat amino acids'. This is further supported by the absence of skin sensitisation response of the read across substance in a human repeat insult patch test following exposure to 5% active substance in human volunteers (CIR, 2013).

A Cosmetic Ingredient Review (CIR) report available for amino acid alkyl amides category reported the below skin sensitisation studies in animals and humans:

Study 2:A study was conducted to determine skin sensitisation potential of read across substance, 'sodium lauroyl silk amino acids', according to local lymph node assay (LLNA) method. In this study, 20% solution of the test substance in butanone was prepared and was applied at three test concentrations: 25, 50 and 100% (leading to final test concentrations of 5, 10 and 20%). The stimulation index (SI) in the study was determined to be 2.61. Under the study conditions, the test substance was determined to be non-sensitising to the skin (CIR, 2013).

Study 3: A study was conducted to determine skin sensitisation potential of read across substance, 'sodium lauroyl glutamate', in 10 female Dunkin-Hartley albino guinea pigs, following exposure to 5% induction and 2.5% challenge test concentrations (in water) according to Modified maximisation test method. Under the study conditions, the test substance was determined to be non-sensitising to the skin. This is further supported by absence of skin sensitisation response of the read across substance in several human repeat insult patch tests following exposure to formulations containing 3-5% active substance in human volunteers (CIR, 2013).

Study 4:A study was conducted to determine skin sensitisation potential of read across substance, 'lauroyl Lysine', in 15 female Dunkin-Hartley albino guinea pigs, following exposure to 50% induction and challenge test concentrations (in olive oil) according to Modified maximisation test method.Under the study conditions, the test substance was determined to be non-sensitising to the skin. This is further supported by absence of skin sensitisation response of the read across substance in a HRIPTs following exposure to formulations (containing 8.36% test substance in blush product and 12.5% test substance in facial powder) under occlusive conditions in 102 and 600 human volunteers respectively (CIR, 2013).

Similar absence of skin sensitisation in HRIPTs with several other alkyl amides was reported in CIR, 2013:

- 2% acetyl hydroxyproline in a plumper gel was determined to be non-sensitising to the skin, when applied to 109 volunteers in a HRIPT under semi-occluded conditions (CIR, 2013).

- 10% acetyl proline in a cream was determined to be non-sensitising to the skin, when applied to 107 volunteers in a HRIPT under semi-occluded conditions (CIR, 2013).

- 1% acetyl tyrosinamide was determined to be non-sensitising to the skin, when applied to a sodium lauryl sulfate pre-treated sites of 26 volunteers in a HRIPT under occlusive conditions. Similar absence of sensitisation response was observed in another HRIPT following application of 2% acetyl tyrosinamide in a plumper gel to 109 volunteers (CIR, 2013).

- 18% solution of disodium capryloyl glutamate (containing 37--41% test substance) was determined to be non-sensitising to the skin, when applied to 20 volunteers in patch test with Finn chambers under occlusive conditions (CIR, 2013).

Further, the CIR Expert Panel, who reviewed the product use, formulation, and safety data of 115 amino acid alkyl amides (which function as skin and hair conditioning agents and as surfactants—cleansing agents in personal care products), concluded that the reviewed amino acid alkyl amides were safe in the present practices of use and concentration in cosmetics, when formulated to be non-irritating. Potassium Lauroyl Wheat Amino Acids, up to 0.7% has CIR safe use history in rinse off cosmetic products (CIR 2013, 2017).

Expert opinions:

Cosmetic Ingredient Review (CIR) Expert Panel: A CIR (2014) report on 'hydrolyzed wheat protein' (HWP) and 'hydrolyzed wheat gluten' (HWG), reported a human repeated insult patch test (HRIPT) study with HWP (MW = 350 Da), which was not found to be a dermal irritant during the induction phase or sensitising during the challenge phase of the study. HWG and HWP are known to have safe use history up to 0.01 and 1.7%, respectively in rinse off cosmetic products. Multiple cases of allergic reactions, including Type 1 immediate hypersensitivity reactions, were reported in individuals who had used personal care products that contained HWP, most of which were to a facial soap in Japan that contained HWP of 40,000-50,000 Da from acid hydrolysis of gluten at high temperatures. Several studies were conducted to characterize the cause, manifestations, and mechanisms of these reactions, including tests of serum IgE binding and reactivity to wheat protein, wheat-protein fractions, and HWP and HWG prepared using acid- and/or enzyme-hydrolysis methods yielding products with varied polypeptide size profiles. From these experiments, it was found that hydrolysates with weight-average MWs < 3000 exhibit no potential to elicit hypersensitivity reactions in sensitized individuals, in contrast to hydrolysates with weight-average MWs >30,000 Da. The experimental results supported the hypothesis that polypeptides with weight-average MWs of 3500 Da or less do not have the potency required to induce Type 1 hypersensitivity. The CIR Panel therefore concluded that data from clinical and laboratory studies was sufficient to demonstrate that these ingredients will not elicit Type 1 immediate hypersensitivity reactions in sensitized individuals, and will not induce sensitization when the polypeptide lengths of the hydrolysates do not exceed 30 amino acids. The Panel concluded that HWG and HWP are safe in cosmetics when formulated to restrict peptides to a weight average of 3500 daltons (Da) or less (CIR, 2014).

Scientific Committee on Consumer Safety (SCCS): The SCCS in Europe published a revised opinion on the safety of 'hydrolysed wheat proteins' in cosmetic products in 2014. In view of the numbers of reported cases of immediate-type contact urticarial and systemic allergic reactions, the overall risk of sensitization to HWP appears to be low, with the exception of an ‘epidemic’ in Japan associated with one particular HWP product used in some brands of soap. Scientific concerns with regard to the use of HWP in cosmetic products include that there is evidence that sensitisation to HWP is via exposure to cosmetics, not via food. Also there are indications that the risk of sensitisation is higher when HWP’s of higher molecular weight are used on the skin, in particular as an ingredient of products that have strong surfactant properties such as soaps and liquid soaps. The SCCS considered the use of hydrolysed wheat proteins safe for consumers in cosmetic products, provided that the maximum molecular weight average of the peptides in hydrolysates is 3.5 kDa (SCCS, 2014).

Basketter and Kimber: Independent Consultants gave their commentary on minimising the risk of allergy to HWP. The authors reviewed the literature on safe use history and occasional allergic reactions reports. They have also reviewed the CIR and SCCS opinion on the HWPs. They have opined that the draft opinion of the SCCS is based on the Japanese Glupearl experience and thereby seeks to limit the risk of HWP allergy in association with cosmetic products. However, they conclude that the optimal approach is to eliminate the hazard at source by imposing an upper limit of molecular weight as proposed by the CIR. Application of a 3.5kD cap on the molecular weight would effectively remove the hazard, such that HWP could continue to be used safely in all cosmetic products.

In general, there is no evidence of any sensitization potential for protein hydrolysates compounds. The protein hydrolyzates are abundantly available in the environment, in food and extensively distributed throughout the human body, which is contradictory to allegation of sensitisation potential. A skin sensitisation test would more or less mimic physiological conditions and thus the result are clearly predictable as “not sensitising”.

Overall, based on weight of evidence information from studies with amino acid alkyl amides and the expert opinions on hydrolysed proteins, the test substance can be considered to be non-sensitising to the skin.

 

References:

1) CIR 2013. Safety Assessment of Amino Acid Alkyl Amides as Used in Cosmetics, Tentative Report for Panel Review, September 20, 2013. Available at https://www.cir-safety.org/sites/default/files/aaaamd092013tent.pdf (accessed in April 2018).

2) SCCS 2014. Opinion on Hydrolysed wheat proteins, SCCS/1534/14 June 18, 2014. Available at ec.europa.eu/health/scientific_committees/consumer_safety/docs/sccs_o_160.pdf (accessed in April 2018).

3) CIR 2017. Safety Assessment of Amino Acid Alkyl Amides as Used in Cosmetics, May 28, 2017.

4) CIR 2014. Safety assessment of hydrolyzed wheat protein and hydrolyzed wheat gluten as used in cosmetics, June 14, 2014. Available at http://www.cir-safety.org/sites/default/files/wheatp062014final.pdf (accessed in April 2018).

5) Basketter D, Kimber I 2014. Hydrolysed wheat proteins: A commentary on minimising the risk of allergy, July 2014 (an independent assessment by 3rd Party skin sensitisation experts for Croda).

Justification for classification or non-classification

Based on weight of evidence information from studies with amino acid alkyl amides and the expert opinions on hydrolysed proteins, the test substance, 'potassium lauroyl wheat amino acids', does not warrant classification for skin sensitisation, according to the EU CLP (Regulation 1272/2008/EC).