Registration Dossier

Administrative data

Description of key information

Fatty acids C16-18, C18 unsat reaction products with tetraethylenepentamine  (FA+TEPA) was found to be sensitising to skin in a GPMT test.

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:
1 Sep 2000 - 25 Sep 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Also see category read across justification document attached to chapter 13
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Read across is done from (source) Fatty acids C16-18, C18 unsat reaction products with tetraethylenepentamine, CAS 68991-84-4 (Tallow-TEPA) to (target) Fatty acids C18 unsat, reaction products with tetraethylenepentamine, CAS 1226892-45-0 (Tall-TEPA)

These amidoamine/imidazolines are made from fatty acid and polyethyleneamines. The manufacturing process is a one-step process with formation of amide and imidazoline structures. To promote imidazoline formation from the amide, the reaction mixture is heated to temperatures above 180ºC. The resulting product therefore is a mixture of the amide structure of the fatty acid and the polyethyleneamine and its imidazoline.
The production of Tallow-TEPA is the same as Tall-TEPA, except for a small difference in chain lengths distribution between tallow derived alkyl chains and tall-oil derived alkyl chains in the fatty acid source used in starting materials. As a consequence, the resulting structures are chemically and structurally completely comparable with only difference in the alkyl chain distribution reflecting the starting material.

Tall oil is basically consisting of C18-unsaturated and some C16 chains, and tallow consists of a mix of C16 (about 25%), C18 (up to 20%) and C18-unsatuarted (up to 50%), and a small percentage other chain lengths.
As consequence, these products are for 50% completely identical, whereas for the other 25% the Tallow-TEPA has a slightly shorter alkyl chain compared to the Tall-TEPA. For all other aspects, the substances are identical. As both have the same chemical structure they both have identical functional groups, show the same chemical reactivity and have very comparable physicochemical properties. Consequently, they share the same mechanism of action. This is similarly applicable to all constituents of these UVCBs.
 
Within a specific structure, the variability of the alkyl chain length is considered to have a possible modifying activity, which is related to modification of the physiological properties of the molecule with increase or shortening of the apolar alkyl chain part. This is suspected to influence aspects related to bioavailability, but not aspects of chemical reactivity and route of metabolism, aspects that influence specific mechanisms of toxicity such as sensitisation and genotoxicity and are more related to the hydrophilic part. As the differences in chain lengths are only very minimal as all substances basically contain C16 and C18 alkyl chains, it seems justified from a toxicological point of view to consider the fatty acid part as similar for all AAI substances.

Specifically for skin sensitisation, the molecular structure with functional groups are essential with respect to the identification of possible skin sensitizing properties. In that respect, Tallow-TEPA and Tall-TEPA are identical.
Related to the (small) difference in average alkyl chain-lengths between the two substances, the Tallow-TEPA with on average a slightly shorter chain-length than the Tall-TEPA, is expected to be slightly better bioavailable (slightly smaller molecule also leading to more moles per unit of weight, slightly higher solubility, slightly lower Kow, and slightly higher vp) and is expected to have (a slightly) higher toxicity profile compared to Tall-TEPA.
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Justification for performing the GPMT instead of the LLNA:

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 GPMT 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 et al., 2008
Comparison of the skin sensitizing potential of unsaturated compounds as assessed by the murine local lymph node assay (LLNA) and the guinea pig maximization test (GPMT).
Food Chem. Toxicol. 46. 1896-1904.
2. Basketter et al., 2009
Application of a weight of evidence approach to assessing discordant sensitisation datasets: Implications for REACH
Regul Toxicol Pharmacol. 55: 90–96
3. Ball et al., 2010
Comparative testing for the identification of skin-sensitizing potentials of nonionic sugar lipid surfactants.
Regul Toxicol Pharmacol. 58(2):301-307.
4. Ball et al., 2011
Evaluating the sensitization 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
Regul Toxicol Pharmacol. 60: 389–400

Additional:
Kreiling et al., 2017, In chemicio, in vitro and in vivo comparison of the skin sensitizing potential of eight unsaturated and one saturated lipid compounds.
Regulatory Toxicology and Pharmacology 90 (2017) 262-276
Specific details on test material used for the study:
name: BASE 136
batch number: AB771
description: thick brown liquid
pH: about 9 (5 % in water)
container: one smoked glass flask
storage conditions: at room temperature and protected from light
composition: see analytical certificate
expiry date: July 2002.
Species:
guinea pig
Strain:
Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
Hartley Cd: (HA) BR, Caesarian obtained, Barrier sustained - Virus Antibody Free (COBS - VAF®)
- Source: Charles River France, 76410 Saint-Aubin-lès-Elbeuf, France.
- Age at study initiation: 1-3 months
- Weight at study initiation: 379 ± 13 g for the males and 374 ± 13 g for the females.
- Housing: individually in polycarbonate cages with stainless steel lid (48 cm x 27 cm x 20 cm) equipped with a polypropylene bottle. Dust-free sawdust was provided as litter (SICSA, 94142 Alfortville, France). Sawdust is analysed by the supplier for composition and contaminant levels.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2°C
- Humidity (%): 30 to 70%
- Air changes (per hr): 12 cycles/hour of filtered, non-recycled air.
- Photoperiod (hrs dark / hrs light): 12 h/12 h

IN-LIFE DATES: From: 1 Sep 2000 To: 25 Sep 2000
Route:
intradermal and epicutaneous
Vehicle:
physiological saline
Concentration / amount:
The dosage form preparation which could pass through a needle and into the dermis had a maximum concentration of 10% (w/w).
Intradermal induction: 0.1%
epicutaneous induction: 5%
epicutaneous challenge: 1%
Route:
epicutaneous, occlusive
Vehicle:
physiological saline
Concentration / amount:
The dosage form preparation which could pass through a needle and into the dermis had a maximum concentration of 10% (w/w).
Intradermal induction: 0.1%
epicutaneous induction: 5%
epicutaneous challenge: 1%
No. of animals per dose:
Control: 5 animals/sex
Treated group: 10 animals /sex
Details on study design:
RANGE FINDING TESTS:
Series of test substance concentrations were tested by intra dermal (0.1, 1, 5, 10% with and without FCA) and epicutaneous route (0.1, 1, 5, 10, 50 100%)
Intradermal: Concentrations of 1% and higher were corrosive. 0.1% resulted to irritation visible during all reported observation day from 24h to 6 days. This concentration was selected for the main study.
Epicutaneous: 5% was the lowest concentration leading to irritation (grade 2: moderate and confluent erythema). This concentration was used for epidermal induction. 1% was without signs of irritation, and was select for challenge concentration.

MAIN STUDY
A. INDUCTION EXPOSURE
-Day 1
The scapular region was clipped and three pairs of intradermal injections (0.1 mL/site) were made in this area as follows:
A) A 1:1 w/w mixture of FCA (Sigma, France) with 0.9% NaCl.
B) The test substance at a 50% concentration in 0.9% NaCl.
C) A 1:1 w/w mixture of the undiluted test substance and FCA.
Note: One of each pair was on each side of the midline and from cranial A) to caudal C).
Controls were treated similarly, but vehicle instead of testsubstance was used.

Day 7
The scapular area between the injection sites was clipped .

Day 8
a pad of filter paper (approximately 8 cm2) was fully-loaded with the test substance at the concentration of 5% (w/w) and was then applied to the interscapular region of the animals of the treated group.
The animals of the control group received an application of the vehicle alone under the same experimental conditions.
The pad was held in place for 48 hours by means of an adhesive hypoallergenic dressing and an adhesive anallergenic waterproof plaster.

B. CHALLENGE EXPOSURE
Day 21: Clipped and shaved each flank.
Day 22: the animals of treated and control groups received an application of the test substance and vehicle. The filter paper of a chamber (Finn Chamber®) was fully-loaded with the test substance at the concentration of 1% (w/w) and was then applied to a clipped area of the skin of the posterior right flank of all animals. The vehicle was applied under the same experimental conditions to the skin of the posterior left flank.
Twenty-four and 48 hours after removal of the dressing of the challenge application, both flanks of the treated and control animals were observed in order to evaluate cutaneous reactions, according to the following scale:
• 0 - no visible change
• 1 - discrete or patchy erythema
• 2 - moderate and confluent erythema
• 3 - intense erythema
Any observed oedema and other lesions were noted..

In life examinations:
At least one a day for clinical signs and mortality
Body weights: Prior to start and at termination of the study.

Positive control substance(s):
yes
Positive control results:
The sensitivity of the experimental technique is regularly assessed using a known moderate sensitizer, MERCAPTOBENZOTHIAZOLE. In a recent study performed under CIT experimental conditions, the strain of guinea pigs used showed a satisfactory sensitization response in 70% animals.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
1% w/w
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Two animals skin injured by dressing, making scoring difficult/impossible; one of these animals also showed crusts.
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 1% w/w. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: Two animals skin injured by dressing, making scoring difficult/impossible; one of these animals also showed crusts..
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
5% w/w
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Two animals skin injured by dressing, making scoring difficult/impossible.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 5% w/w. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: Two animals skin injured by dressing, making scoring difficult/impossible..
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
5%
No. with + reactions:
19
Total no. in group:
19
Clinical observations:
One animal died study day 11 (before challenge). Dryness of skin and/or crusts observed in most animals.
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 5%. No with. + reactions: 19.0. Total no. in groups: 19.0. Clinical observations: One animal died study day 11 (before challenge). Dryness of skin and/or crusts observed in most animals..
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
5% w/w
No. with + reactions:
19
Total no. in group:
19
Clinical observations:
Dryness of skin and/or crusts observed in all animals.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 5% w/w. No with. + reactions: 19.0. Total no. in groups: 19.0. Clinical observations: Dryness of skin and/or crusts observed in all animals..
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
20% (w/w)
No. with + reactions:
7
Total no. in group:
10
Clinical observations:
dryness ofthe skin & oedema
Remarks on result:
positive indication of skin sensitisation
Interpretation of results:
sensitising
Remarks:
Migrated information
Conclusions:
Reactions were observed in all animals of treatment group and none in the control. Cutaneous effects were attributed to delayed contact hypersenisitivity.
Executive summary:

The potential of the test substance BASE 136 (batch No. AB77l) to induce delayed contact hypersensitivity was evaluated in guinea pigs according to the maximization method of Magnusson and Kligman and to OECD (No. 406, 17th July 1992) and EC (96/S4/EEC, B.6, 30 July 1996) guidelines.

The study was conducted in compliance with the principles of Good Laboratory Practice Regulations.

 

 

Thirty guinea pigs were allocated to two groups: a control group of five males and five females and a treated group of ten males and ten females.

 

On day 1, three pairs of intradermal injections were performed in the interscapular region of all animals:

- Freund's complete adjuvant (FCA) diluted at 50% with 0.9% NaCl (both groups),

- test substance at the chosen concentration in the chosen vehicle (treated group) or vehicle alone (control group),

- test substance at the chosen concentration in a mixture FCA/0.9% NaCl 50/50 (treated group) or vehicle at the concentration of SO% in a mixture FCAl/0.9% NaCl 50/50 (control group).

 

On day 8, the test substance (treated group) or the vehicle (control group) was applied topically to the same test site, which was then covered by an occlusive dressing for 48 hours.

 

On day 22, all animals of the treated and control groups were challenged by a cutaneous application of the test substance to the right flank. The left flank served as control and received the vehicle only. Test substance and vehicle were maintained under an occlusive dressing for 24 hours.

Skin reactions were evaluated approximately 24 and 48 hours after removal of the dressing.

 

Test substance concentrations were as follows:

 

Induction (treated group)

-       intradermal injections (day 1): BASE 136 at the concentration of 0.1% (w/w) in sterile isotonic saline solution (0.9% NaCl),

-       topical application (day 8): BASE 136 at the concentration of 5% (w/w) in sterile isotonic saline solution (0.9% NaCl).

 

Challenge (all groups)

-       topical application (day 22): BASE 136 at the concentration of 1% (w/w) in sterile isotonic saline solution (0.9% NaCl).

 

At the end of the study, animals were killed without examination of internal organs.

Skin samples were taken from the challenge application sites of all the animals.

No histological examination was performed.

 

Results

No clinical signs and no deaths related to treatment were noted during the study.

After the challenge application, no relevant cutaneous reactions were observed in the animals of the control group.

In the treated group, a discrete or moderate erythema was noted in all animals. An oedema was recorded in 11/19 animals. Dryness of the skin and/or crusts were observed all animals. The observed cutaneous were attributed to delayed contact hypersensitivity.

 

Conclusion

Under our experimental conditions and according to the maximization method of Magnusson and Kligman, the test substance BASE 136 (batch No. AB771) induces delayed contact hypersensitivity in 19/19 (100%) guinea pigs.

According to the classification criteria laid down in Directive 93/21/EEC (27th April 1993) adapting to technical progress for the eighteenth time Council Directive 67/548/EEC, the test substance should be considered as a skin sensitizer.

These results are valid also for tall oil + TEPA, as Fatty acids C16-18, C18 unsat and Tall oil are only marginally differing in chain lengths distributions, and it is considered that both their reaction products with TEPA are principally the same when evaluating for sensitisation.

These results lead to classification according to CLP (ATP 2): Skin sensitiser Cat.1A (GPMT ≥ 30% positive at ≤ 0.1% i.d. induction)

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

Fatty acids C16-18, C18 unsat reaction products with tetraethylenepentamine was found to be sensitising to skin in a GPMT test. These results are valid also for tall oil + TEPA, as Fatty acids C16-18, C18 unsat and Tall oil are only marginally differing in chain lengths distributions, and it is considered that both their reaction products with TEPA are principally the same when evaluating for sensitisation.

The GPMT study was performed according to OECD 406 andGLP compliant. Thirty guinea pigs were allocated to two groups: a control group of five males and five females and a treated group of ten males and ten females.Induction of the treated group involved intra dermal injection on day 1 of concentration of 0.1% (w/w), and topical application of 5% (w/w) in sterile isotonic saline solution on day 8. The challenge on day 22 involved topical application of 1% (w/w). Positive skin reactions were observed in all animals of the treatment group, and in none of the animals of the control group.

The following information is available on sensitising properties of AAI substances: 

TO + DETA

Sensitising: 40% positive reaction in GPMT, i.d. induction 0.1%

TO + DETA 

Sensitising: LLNA: EC3: 0.3%.

C16-18, C18 unsat + TEPA

Sensitising: 100% positive reaction in GPMT, i.d. induction 0.1%

TO + Poly(Amide)

 Sensitising: 100% positive reaction in GPMT, i.d. induction 0.2%

Based on the available data, all substances in the AAI group should be considered sensitising to skin.

Classification according to CLP (ATP 2) skin sensitiser: Cat.1A (GPMT ≥ 30% positive at ≤ 0.1% i.d. induction)

 

For corrosive substances, the use of protective gloves and other equipment, such as face shields, aprons and good work practices are mandatory. As a result, direct dermal contact is very unlikely.

Lack of exposures explains that although the substance can be considered a strong sensitiser based on the results from both studies, the producers are not aware of reported incidences of sensitisation among own employees or from customers.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available
Additional information:

There is no information on respiratory sensitisation, but Fatty acids C16-18, C18 unsat reaction products with tetraethylenepentamine was sensitising to skin.

The likelihood for exposure via inhalation and thus becoming sensitised to AAI, is very low, based on the high boiling point (> 300 °C) and very low vapour pressure (0.00017 mPa at 25°C for DETA based AAI, which is considered to have the highest vapour pressure within the AAI). The potential for inhalation is not significant to justify this study. Furthermore, as the substance is classified as corrosive, such testing should normally not be conducted.

Justification for classification or non-classification

Various studies with different AAI indicate that these substances can cause dermal sensitisation. Fatty acids C16-18, C18 unsat reaction products with tetraethylenepentamine was sensitising to skin in a GPMT. Based on the results of this study FA+TEPA should be classified as dermal sensitiser Cat.1A for CLP

 

The actual risk of sensitisation is probably low, as AAI are corrosive to skin and consequently exposure will be low due to necessary protective measures to limit dermal exposure.

Also the likelihood for exposure via inhalation and thus becoming sensitised to AAI, is very low, based on the high boiling point (> 300 °C) and very low vapour pressure (0.00017 mPa at 25°C for DETA based AAI).