Registration Dossier

Administrative data

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015-12-07 to 2016-04-13
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
(Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany)
Test type:
standard acute method
Limit test:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
2,2'-(octylimino)bisethanol
EC Number:
239-555-0
EC Name:
2,2'-(octylimino)bisethanol
Cas Number:
15520-05-5
Molecular formula:
C12H27NO2
IUPAC Name:
2-[(2-hydroxyethyl)(octyl)amino]ethan-1-ol
Test material form:
other: liquid
Details on test material:
Name: Genamin 3920
CAS No.: 15520-05-5
Molecular Weight of Base Form: 217.3483
Chemical Name: 2,2’-(Octylimino)bisethanol
Physical State at RT: liquid
Colour: clear
pH: 11.6 (23 °C, water:ethanol 1:1)

Storage Conditions: 2 – 8 °C, protected from light

Safety Precautions: The routine hygienic procedures are sufficient to assure personnel health and safety.

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Housing and Feeding Conditions
- Full barrier in an air-conditioned room
- Temperature: 22 +/- 3 °C
- Relative humidity: 55 +/- 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: 10 x / hour
- Free access to Altromin 1324 maintenance diet for rats and mice (lot no. 02102150820)
- Free access to tap water, sulphur acidified to a pH value of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals)
- The animals were kept individually in IVC cages, type III H, polysulphone cages on Altromin saw fibre bedding (lot no. 0446 )
- Certificates of food, water and bedding are filed for two years at BSL Munich and afterwards archived at Eurofins Munich
- Adequate acclimatisation period (at least five days) under laboratory conditions

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
Preparation of the Animals:
The animals were marked for individual identification by tail painting.
Approximately 25 hours before the test, the fur was removed from the dorsal area of the trunk using an electric clipper. Care was taken to avoid
abrading the skin, and only animals with healthy intact skin were used.
No less than 10% of the body surface was cleared for the application.
Prior to the application a detailed clinical observation was made of all animals. Only healthy animals were used.

Application:
The test item was applied at a single dose, uniformly over an area which was approximately 10% of the total body surface.
The test item was held in contact with the skin by a dressing throughout a 24-hour period. The dressing consisted of a gauze-dressing and
non-irritating tape and was fixed with an additional dressing in a suitable manner.

Duration of exposure:
The test item was held in contact with the skin throughout a 24-hour period. At the end of the exposure period the residual test item was not
removed.
Doses:
The test item was applied at a single dose of 2000 mg/kg body weight to each animal.
No. of animals per sex per dose:
5 male and 5 female
Control animals:
not required
Details on study design:
Observation period:
All animals were observed for 14 days after dosing

Weight Assessment:
The animals were weighed on day 1 (prior to the application) and on days 8 and 15.

Clinical Examination:
careful clinical examination was made several times on the day of dosing (at least once during the first 30 minutes and with special attention given
during the first 4 hours post-dose). As soon as symptoms were noticed they were recorded. Thereafter, the animals were observed for clinical signs once daily until the end of the observation period. All abnormalities were recorded.
Cageside observations included changes in the skin and fur, eyes and mucous membranes. Also respiratory, circulatory, autonomic and central
nervous systems and somatomotor activity and behaviour pattern were examined. Attention was directed to observations of tremors, convulsions,
salivation, diarrhoea, lethargy, sleep and coma.

Pathology:
At the end of the observation period the animals were sacrificed with an overdosage of pentobarbital injected intraperitoneally (Narcoren®, Merial;
lot no.: 250055; expiry date: 31/05/2018) at a dosage of 250-400 mg/kg bw. All animals were subjected to gross necropsy and examined
macroscopically for gross pathological changes. In absence of gross pathological changes no tissues were preserved for a possible histopathologicalevaluation.

Evaluation of Results:
Individual reactions of each animal were recorded at each time of observation.
Toxic response data were recorded by sex and dose level.
Nature, severity and duration of clinical observations were described.
The body weight changes were summarised in a tabular form.
Necropsy findings were described.
With few exceptions, data were captured using the validated departmental computerised system E WorkBook (version 9.4.0, ID Business Solutions Ltd.)
Statistics:
According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the
results is not regarded as necessary.

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality was observed
Clinical signs:
No treatment-related effects were observed.
Body weight:
A slight weight loss was recorded for 4 out of 5 female animals during the first week and for 1 out of 5 female animals during the second week.
The effects on weight development might be secondary to the dressing, and toxicological relevance of this finding cannot clearly be concluded.
The male animals showed weight gain during the first and the second weeks of observation.
Gross pathology:
On the skin and in the subcutis either scab or scab and one or more red or dark foci were observed in all animals.
Other findings:
Erythema up to grade 2 and oedema up to grade 2 were observed in all animals. Either necrosis or eschar or both were observed in all animals.
The signs of irritation were not reversible within the observation period.

Any other information on results incl. tables

Clinical Signs of Systemic Toxicity - Individual Data - Males

Animal No.

Dose (mg/kg bw)

Timepoint

Observations

Comment

21

2000

(0min – 240min)

Nsf

-

21

2000

(2d – 15d)

Nsf

Local findings

22

2000

(0min – 240min)

Nsf

-

22

2000

(2d- 5d)

Nsf

Local findings

22

2000

(6d – 12d)

Nsf

-

22

2000

(13d – 15d)

Nsf

Local findings

23

2000

(0min – 6d)

Nsf

-

23

2000

(7d)

Nsf

Local findings

23

2000

(8d – 12d)

Nsf

-

23

2000

(13d – 15d)

Nsf

Local findings

24

2000

(0min – 6d)

Nsf

-

24

2000

(7d – 15d)

Nsf

Local findings

25

2000

(0min – 240min)

Nsf

-

25

2000

(2d – 15d)

Nsf

Local findings

min = minute(s); h = hour(s); d = day(s), bw = body weight;nsf = no specific findings Clinical Signs of Systemic Toxicity - Individual Data - Females

Animal No.

Dose (mg/kg bw)

Timepoint

Observations

Comment

26

2000

(0min – 240min)

Nsf

-

26

2000

(2d – 15d)

Nsf

Local findings

27

2000

(0min- 12d)

Nsf

-

27

2000

(13d – 15d)

Nsf

Local findings

28

2000

(0min – 240min)

Nsf

-

28

2000

(2d – 15d)

Nsf

Local findings

29

2000

(0min – 240min)

Nsf

-

29

2000

(2d – 15d)

Nsf

Local findings

30

2000

(0min – 240 min)

Nsf

-

30

2000

(2d – 15d)

Nsf

Local findings

min = minute(s); h = hour(s); d = day(s), bw = body weight;nsf = no specific findings

Skin Irritation at Application Site – Individual Data – Males

Animal No. Timepoint Oedema/ Local Comment
Erythema Skin Findings
21 (2d – 5d) 0/0      Necrosis Black focus at application site approx. 2cmx2cm
21 (6d – 8d) 1/2 Necrosis Black focus approx. 3cmx4cm
21 (9d -12d) 1/1 Necrosis Black focus approx. 3cmx4cm
21 (13d – 15d) 0/1      Necrosis Eschar peeling off, black focus approx. ø 1cm
22 (2d) 0/0      Necrosis Black focus at application site approx. 2cmx2cm
22 (3d – 4d) 0/0      Necrosis Black focus at application site approx. ø 0.5cm
22 (5d) 1/1 Necrosis Black focus at application site approx. ø 0.5cm
22 (6d) 2/2 Nsf -
22 (7d) 1/2 Nsf -
22 (8d – 12d) 0/1      Nsf -
22 (13d - 14d) 0/0      Eschar 3 foci ø 0.5cm
22 (15d) 0/0      Eschar 2 foci ø 0.3cm
23 (2d – 4d) 0/0      Nsf -
23 (5d) 1/1 Nsf -
23 (6d) 2/2 Nsf -
23 (7d) 2/2 Necrosis Black focus approx. 1.5cmx1.5cm
23 (8d – 12d) 0/1      Nsf -
23 (13d) 0/0      Eschar 1 focus ø 0.5 cm
23 (14d – 15d) 0/0      Eschar 1 focus ø 0.3 cm
24 (2d – 4d) 0/0      Nsf -
24 (5d) 1/1 Nsf -
24 (6d) 2/2 Nsf -
24 (7d) 2/2 Necrosis Black focus approx. 2cmx2cm
24 (8d) 1/1 Necrosis Black focus approx. 1cmx1cm
24 (9d – 12d) 0/1      Necrosis Black focus approx. 1cmx1cm
24 (13d) 0/0      Eschar 1 focus ø 0.5cm
24 (14d – 15d) 0/0      Eschar 1 focus ø 0.3cm
25 (2d) 0/0      Necrosis Black focus at application site approx. 2cmx2cm
25 (3d – 4d) 0/0      Necrosis Black focus at application site approx. 1cm
25 (5d) 1/1 Necrosis Black focus at application site approx. 1cm
25 (6d) 1/2 Necrosis Black focus approx. 1cmx0.5cm
25 (7d) 1/2 Necrosis Black focus approx. 2cmx2cm
25 (8d) 1/1 Necrosis Black focus approx. 1.5cmx1.5cm
25 (9d – 12d) 0/1      Necrosis Black focus approx. 1.5cmx1.5cm
25 (13d – 15d) 0/0      Eschar 2 foci ø 0.3cm
E = erythema; O = oedema; 0, 1, 2, 3, 4 = scoring system laid down in OECD Guideline 404 Skin Irritation at Application Site - Individual Data - Females
Animal No. Timepoint Oedema/ Local Comment
Erythema Skin Findings
26 (2d – 4d) 0/0      Necrosis  Black focus at application site approx. 2cmx2cm
26 (5d) 2/2 Necrosis  Black focus at application site approx. 2cmx2cm
26 (6d – 12d) 2/2 Necrosis  Black focus approx. 3cmx4cm
26 (13d – 15d) 0/1      Necrosis  Black focus approx. 2cmx2cm, eschar peeling off 
27 (2d – 3d) 0/0      nsf  -
27 (4d) 0/2      nsf  -
27 (5d) 1/2 nsf  -
27 (6d – 7d) 2/2 nsf  -
27 (8d – 12d) 0/1      nsf  -
27 (13d – 15d) 0/0      Eschar 1 focus approx. ø 0.4cm
28 (2d – 4d) 0/0      Necrosis  Black focus at application site approx. 2cmx2cm
28 (5d) 1/2 Necrosis  Black focus at application site approx. 2cm.x2cm
28 (6d – 7d) 2/2 Necrosis  Black focus approx. 2cmx2cm
28 (8d – 12d) 1/1 Necrosis  Black focus approx. 2cmx2cm
28 (13d – 15d) 0/1      Necrosis  Black focus approx. 2cmx2cm
29 (2d – 3d) 0/0      Necrosis  Black focus at application site approx. 2cmx2cm
29 (4d) 0/1      Necrosis  Black focus at application site approx. 2cmx2cm
29 (5d) 0/2      Necrosis  Black focus at application site approx. 2cmx2cm
29 (6d) 1/2 Necrosis  Black focus approx. 2cmx2cm
29 (7d) 1/2 Necrosis  Black focus approx. 2.5cmx2.5cm
29 (8d) 1/1 Necrosis  Black focus approx. 2.5cmx2.5cm
29 (9d – 12d) 0/1      Necrosis  Black focus approx. 2.5cmx2.5cm
29 (13d) 0/0      Necrosis  Black focus approx. 2.5cmx2.5cm
29 (14d – 15d) 0/0      Eschar Eschar peeling off, approx. ø 1cm
30 (2d – 4d) 0/0      Necrosis  Black focus at application site approx. 2cmx2cm
30 (5d – 6d) 1/2 Necrosis  Black focus at application site approx. 2cmx2cm
30 (7d) 2/2 Necrosis  Black focus approx. 2.5cmx2.5cm
30 (8d – 12d) 1/1 Necrosis  Black focus approx. 2.5cmx2.5cm
30 (13d – 14d) 0/1      Necrosis  Black focus approx. 1cmx1cm, eschar peeling off
30 (15d) 0/0      Eschar  Eschar peeling off, approx. ø 1cm

E = erythema; O = oedema; 0, 1, 2, 3, 4 = scoring system laid down in OECD Guideline 404;
nsf = no specific findings

Absolute Body Weights in g and Body Weight Gain in %

Test Group Animal No. Dose (mg/kg bw) Body Weight (g) on Day BW gain in Comparison to Day 1 (%)
Day 1 Day 8 Day 15 Day 15
Males 21 2000 243 252 258 6
22 241 257 278 15
23 244 257 280 15
24 267 283 321 20
25 241 257 282 17
Females 26 2000 231 227 212 -8
27 232 228 235 1
28 225 221 227 1
29 230 237 243 6
30 220 218 225 2

bw = body weight

Macroscopic Findings - Individual Data - Males and Females

Test Group Animal No. Dose (mg/kg bw) Organ Macroscopic Findings
Males 21 2000 Skin Scab 1cm
Subcutis Focus red 1cm
22 2000 Skin Scab 2cmx0.3cm
23 2000 Skin Scab 0.3cm
24 2000 Skin Scab 0.3cm
25 2000 Skin Scab 2cmx0.3cm
Females 26 2000 Skin Scab 4cm
Subcutis Focus dark 1cm;
focus red 3cm
27 2000 Skin Scab 0.4cm
Subcutis Focus red 0.2cm
28 2000 Skin Scab 4cm
Subcutis Focus red 1cm
29 2000 Skin Scab 1cm
Subcutis Focus red 1cm
30 2000 Skin Scab 1cm
Subcutis Focus red 1cm

nsf = no specific findings

LD50

Test Group Dose (mg/kg bw) Number of Animals Number of Intercurrent Deaths LD50Cut-Off             (mg/kg bw)
Males 2000 5 0 > 2000
Females 2000 5 0

bw = body weight

Applicant's summary and conclusion

Interpretation of results:
relatively harmless
Remarks:
Migrated information severely irritating but no systemic toxicity Criteria used for interpretation of results: EU
Conclusions:
Under the conditions of the present study, single dermal application of the test item Genamin 3920 to rats at a dose of 2000 mg/kg body weight was
associated with no mortality and no signs of systemic toxicity but with signs of irritation/corrosion at the application site which were not fully
reversible within the observation period.
The dermal LD50 was determined to be > 2000 mg Genamin 3920 / kg body weight.
Executive summary:

Summary Results

LD50:                                      > 2000 mg/kg bw

Species/strain:                      WISTAR Crl: WI(Han) rats

Vehicle (moistening):             no vehicle used

Number of animals:                5 male and 5 female

Duration of exposure:           24 hours

Method:                                OECD 402, EC 440/2008, Method B.3

Results

Test Group

Animal No.

Dose (mg/kg bw)

Number of Animals

Number of Intercurrent Deaths

Males

21 - 25

2000

5

0

Females

26 - 30

2000

5

0

bw = body weight

Signs of systemic toxicity related to dose level used, time of onset and duration:

No treatment-related effects were observed.

Effect on organs (related to dose level):

On the skin and in the subcutis either scab or scab and one or more red or dark foci were observed in all animals.

Signs of irritation:

Erythema up to grade 2 and oedema up to grade 2 were observed in all animals.Either necrosis or eschar or both were observed in all animals.

The signs of irritation were not reversible within the observation period.

Conclusion

Under the conditions of the present study, single dermal application of the test itemGenamin 3920to rats at a dose of 2000 mg/kg body weight was associated with no mortality and no signs of systemic toxicity but with signs of irritation/corrosion at the application site which were not fully reversible within the observation period.

The dermal LD50was determined to be > 2000 mgGenamin 3920/ kg body weight.