Registration Dossier

Administrative data

Description of key information

oral (OECD 401): LD50mouse = 2640 mg/kg bw (neat substance)

dermal (OECD 402 limit test): LD50rat > 2000 mg/kg bw (31% a.i.), equivalent to 620 mg/kg bw (neat substance)

inhalation: Data waiving

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study meets generally accepted scientific principles, acceptable for assessment. Only very limited documentation, only 5 days observation period.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Deviations:
yes
Remarks:
Only very limited documentation, only 5 days observation period
GLP compliance:
no
Test type:
standard acute method
Limit test:
no
Species:
mouse
Strain:
CF-1
Sex:
not specified
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River
- Weight at study initiation: 18 - 21 g
- Fasting period before study: no data
- Housing: 5/cage
- Diet (e.g. ad libitum): Lab Blox, ad libitum
- Water (e.g. ad libitum): ad libitum
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 10 % (test material was diluted 1:3 in water just prior to the test)
Doses:
6670, 8350, 10000 mg/kg bw
No. of animals per sex per dose:
10 (sex not specified)
Control animals:
no
Details on study design:
- Duration of observation period following administration: 5 days
- Necropsy of survivors performed: no data
Statistics:
Oral LD50 was calculated according to the method of Litchfield and Wilcoxon (1949).
Sex:
not specified
Dose descriptor:
LD50
Effect level:
2 640 mg/kg bw
Based on:
act. ingr.
95% CL:
2 310 - 3 000
Remarks on result:
other: recalculated to neat substance
Sex:
not specified
Dose descriptor:
LD50
Effect level:
8 800 mg/kg bw
Based on:
test mat.
95% CL:
7 700 - 10 000
Mortality:
10000 mg/kg: 7/10
8350 mg/kg: 4/10
6670 mg/kg: 1/10
Clinical signs:
not reported
Body weight:
not reported
Gross pathology:
not reported

According to the criteria of EU Directive 67/548/EEC and the criteria of Regulation (EC) No 1272/2008 the test substance does not have to be classified for acute oral toxicity.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
2 640 mg/kg bw
Quality of whole database:
The available information comprises an adequate and reliable study (reliability 2) for a reference substance with similar structure and intrinsic properties and a supporting study for the registration substance itself with slight qualitative deficiencies. Taken together, the information from these independent sources is consistent and provides sufficient quality for hazard assessment leading to an endpoint conclusion in accordance with Annex XI, 1.2, of Regulation (EC) No 1907/2006. Therefore, the available information as a whole is sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1987-07-27 to 1987-08-10
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across from a guideline study
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: CD (Crl:COBS CD(SD)BR)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River U.K. Limited, Margate, Kent, England
- Age at study initiation: 7 - 10 weeks
- Weight at study initiation: 200 to 232 g
- Fasting period before study: no
- Housing: individually housing in metal cages with wire mesh floors
- Diet: ad libitum, standard laboratory rodent diet "Labsure LAD 1"
- Water: ad libitum
- Acclimation period: 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 23
- Humidity (%): 66
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
One day prior to treatment hair was removed from the dorsolumbar region of each rat with electric clippers exposing an area equivalent to 10% of the total body surface. No shaving or chemical depilation was used.

The test substance was applied by spreading it evenly over the prepared skin. Total volume applied was 1.92 ml/kg bw. Test substance was applied in the original state (aqueous solution, a.i. 31 %) as delivered by the sponsor. The treated area was then promptly covered with gauze which was held in place with an impermeable dressing encircled firmly around the trunk.

At the end of the 24-hours exposure period, the dressings were carefully removed and the treated area of skin decontaminated by washing in warm (30°-40°C) water and blotting dry with absorbent paper.
Duration of exposure:
24 hours
Doses:
2000 mg/kg bw (based on product),
No. of animals per sex per dose:
5 males
5 females
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 d
- Body weights were recorded on Day 1 (day of dosing), 8 and 15
- Clinical observation: Animals were observed soon after dosing and at frequent intervals for the remainder of Day 1. On subsequent days the animals were observed once in the morning and again at the end of the experimental day. This latter observation was at approximately 16.30 hours on week days or 11.30 hours on Saturday and Sunday. Clinical signs were recorded at each observation. The nature, severity, approximate time of
onset and duration of each toxic sign.
- Necropsy of the survivors performed: yes
- Other examinations performed: skin reactions - The treated areas of skin were examined daily for signs of dermal irritation and assessed according to the following arbitrary scoring system.
Erythema and eschar formation:
No erythema = score 0
Slight erythema = score 1
Well-defined erythema = score 2
Moderate to severe erythema = score 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth) = score 4
Oedema formation:
No oedema = score 0
Slight oedema = score 1
Well-defined oedema (area well-defined by definite raising) = score 2
Moderate oedema (raised approximately 1 millimetre) = score 3
Severe oedema (raised more than 1 millimetre and extending beyond the area of exposure) = score 4
A separate record was kept of dermal changes other than erythema and oedema.
Sex:
male/female
Dose descriptor:
LD0
Effect level:
> 2 000 mg/kg bw
Remarks on result:
other: based on product
Sex:
male/female
Dose descriptor:
LD0
Effect level:
> 620 mg/kg bw
Remarks on result:
other: based on a.i.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Remarks on result:
other: based on product; mortality 0/10
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 620 mg/kg bw
Remarks on result:
other: based on a.i.; mortality 0/10
Mortality:
0/10
Clinical signs:
- Application site: Sites of application of the test substance showed slight or well-defined erythema in all animals on Day 2 and/or 3. On Day 4 and Day 5, slight erythema were seen in three male and four female rats, sloughing in two male rats and one female rat and hyperkeratinisation in three female rats. All skin reactions were completely reversible by Day 6 in all animals.
Slough or hyperkeratinisation affected the treated skin of 4, 5, 6, 8, 9, 10 rats on Days 4 and 5 only.

- systemic: There were no clinical signs of systemic reaction to treatment.
Body weight:
Slightly low bodyweight gains were recorded for three females on Day 8.
All other rats achieved anticipated bodyweight gains throughout the study.
Gross pathology:
Terminal autopsy findings were normal.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: other: CLP, EU GHS (Regulation (EC) No 1272/2008)
Conclusions:
The acute lethal dermal dose to rats was found to be > 2000 mg/kg bodyweight (based on product).
Executive summary:

In an acute dermal toxicity study according to EU Method B.3 and OECD Guideline 402, 5 male and 5 female CD rats (Crl:COBS CD(SD)BR) were dermally exposed to Coco AAPB (a.i 31 %) as delivered by the sponsor for 24 hours to 10% of total body surface at a dose of 2000 mg/kg bw (limit test). Test sites were covered with an occlusive dressing. After 24 hours, the test sites were rinsed with warm water. Animals then were observed for 14 days after dosing.

There were no clinical signs of systemic reaction to treatment. Sites of application of the test substance showed slight or well-defined erythema in all animals on Day 2 and/or 3. On Day 4 and Day 5, slight erythema were seen in three male and four female rats, sloughing in two male rats and one female rat and hyperkeratinisation in three female rats. All skin reactions were completely reversible by Day 6 in all animals. Slightly low bodyweight gains were recorded for three females on Day 8. All other rats achieved anticipated bodyweight gains throughout the study. Terminal autopsy findings were normal.

Dermal LD0 Combined: 2000 mg/kg bw

Dermal LD50 Combined: > 2000 mg/kg bw

LD0 and LD50 determined refer to the test substance as delivered by the sponsor. Amount of active ingredient in test substance is 31 %. Therefore the calculated oral LD0and LD50 combined referring to 100 % active substance is 620 and > 620 mg/kg bw, respectively.

Coco AAPB (a.i. 31 %) is of low toxicity based on the LD50 in males and females.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw
Quality of whole database:
The available information comprises an adequate and reliable study (reliability 2) for a reference substance with similar structure and intrinsic properties. The available study is sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Additional information

Two reliable studies are available addressing acute oral toxicity, the first one performed with an undiluted aqueous formulation of CAS 66455-29-6 (with which Betaines, C12-14 (even numbered)-alkyldimethyl was previously referred to) in rats, but lacking specification of the active ingredient content (Huntingdon Research Centre, 1969), and the second one with an undiluted aqueous solution containing 30% a.i. of CAS 68424-94-2 in mice (Levenstein, 1979). The most sensitive one was chosen as key study, which was done with CAS 68424-94-2, demonstrating a LD50 of 2640 mg/kg bw, recalculated to neat substance, in mice. With CAS 66455-29-6 a LD50 of 3202.5 mg/kg bw was demonstrated in rats for a not sufficiently specified aqueous formulation. The mortality observed in this study had to be attributed to local irritant effects of the test substance, manifesting in form of gastro-intestinal inflammation in the deceased animals only, rather than systemic toxicity. Both values were in the same range of magnitude. Therefore, 2640 mg/kg bw was adopted as oral LD50 as the most sensitive value. Those values are in line with an approximate LD100 of 7500 mg/kg bw reported for CAS 683-10-3 by a supporting study (Haskell Laboratory, 1963). Further information for that substance is available from a secondary source reporting a result derived from the acute toxic class method (Kashima Laboratory, 2005). In this study a classification into acute toxicity class 4 (LD50 >300 - 2000 mg/kg bw) was derived. The original data source was not available, therefore it was not chosen for assessment. Additionally, an LD50 of 71 mg/kg bw is reported in a publication (Ridout et al., 1991). Although being the lowest value in the range this publication was not chosen for assessment. First of all it suffers from extremely limited documentation, as even dose levels were not reported, and secondly the value is extraordinarily low in comparison with any other available data, which lacks a logical explanation. Therefore, this publication was not considered suitable for assessment.

As there was no data of sufficient quality available adressing acute dermal toxicity for the substance itself, read-across data from a structural analogue (cocamidopropyl betaine, CAS 61789-40-0) was used (Kao, 1987). This study was designed as a limit test with application of 2000 mg/kg bw of an aqueous solution with 31% active ingredient (commercial product, as supplied) to the skin of rats for a period of 24 hours under occlusive conditions. No mortality occurred, and no clinical signs were observed within the 14-day observation period. There were no unusual findings at terminal autopsy, the only observations were signs of slight to well-defined local skin irritation at the application sites. Therefore, an LD50 > 2000 mg/kg bw was established for the commercial product with 31% active ingredient, the dermal LD50 for 100% active substance was calculated to be > 620 mg/kg bw.

A dermal LD50 of 1300 mg/kg bw was reported for the substance itself in the publication already mentioned for acute oral toxicity testing (Ridout et al., 1991). Although providing the lowest value in comparison with any other available data, this publication was not considered for assessment. The same deficiencies valid for oral toxicity testing within this publication are also valid for testing of dermal toxicity. First of all it suffers from extremely limited documentation, as only 2 dose levels were tested which were even not reported, and secondly the LD50 value for dermal toxicity is lower than that established for oral toxicity for the betaines in reliable studies. This is not feasible, as the same authors demonstrated plausibly within the same publication a dermal absorption of 46.5% through mouse skin within 24 hours in vitro. Therefore, dermal LD50 is expected to be higher than the oral one. Additionally, based on common scientific experience, it is justified to anticipate a lower penetration rate through intact skin in vivo. These discrepancies were not addressed and lack a logical explanation. Therefore, this publication was not considered suitable for assessment.

However, although the read-across study conducted with cocamidopropyl betaine does not cover the limit dose requested by OECD guideline 402, carrying out additional acute dermal studies for the pure substances is scientifically not justified as dermal absorption for human skin in vivo was shown to be negligible (max. 0.4% within 30 min), and acute oral toxicity was demonstrated to be very low, either. Therefore, anticipation of higher toxicity via the dermal route than via the oral route is scientifically unjustified, and a classification for acute dermal toxicity is not to be expected. Additionally, the substance is only marketed in form of aqueous solutions with active substance concentrations not exceeding 31%, which further reduces the possibility of uptake of eventually toxic doses via the skin.

Inhalation:

According to Regulation (EC) No 1907/2006, 8.5.2, column 2, administration by inhalation is not required. The test substance has a low vapour pressure, exposure to aerosols, particles or droplets is unlikely, and appropriate RMMs are already implemented. Data for the oral and dermal route are available, additional inhalation testing would neither improve risk assessment nor safety of applications.

Justification for selection of acute toxicity – oral endpoint

Hazard assessment is conducted by means of read-across from a structural analogue, although a study conducted with the registration substance itself is available.  All available studies are adequate and reliable based on the identified similarities in structure and intrinsic properties and overall assessment of quality, duration and dose descriptor level (refer to the endpoint discussion for further details), but the selected study is the most sensitive one, has the lowest LD50 and a reproducible documentation of the applied dose.

Justification for selection of acute toxicity – dermal endpoint

Hazard assessment is conducted by means of read-across based on a read-across from a structural analogue because no information is available for the substance itself. The selected study is adequate and reliable based on the identified similarities in structure and intrinsic properties among the target and the source substance and overall assessment of quality, duration and dose descriptor level (refer to the endpoint discussion for further details).

Justification for classification or non-classification

Based on the available data for substances structurally closely related to Betaines, C12-14 (even numbered)-alkyldimethyl according to the criteria of Regulation (EC) No 1907/2006, Annex XI, section 1.5 an oral LD50 of 2640 mg/kg bw determined in mice has been adopted; therefore, Betaines, C12-14 (even numbered)-alkyldimethyl does not have to be classified for acute oral toxicity according to EU Directive 67/548/EEC and Regulation (EC) No 1272/2008. The dermal LD50 for a neat structural analogue was determined to be > 620 mg/kg bw (limit test with 2000 mg/kg bw of 31% aqueous test substance solution). Nevertheless, considering the dermal absorption data it is justified to assume that toxicity via the dermal route is not higher than that via the oral route; therefore, a classification of Betaines, C12-14 (even numbered)-alkyldimethyl for acute dermal toxicity is not justified, either.