Registration Dossier

Administrative data

Description of key information

Oral (OECD 423), rat: LD50 cut-off = 5000 mg/kg bw

Inhalation (OCED 436), rat, 4 h exposure: LC50 cut-off = 3.81 mg/L

Dermal (OECD 402), rat: LD50 > 2000 mg/kg bw

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017-01-17 to 2017-04-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
December 17, 2001
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
May 30, 2008
GLP compliance:
yes (incl. certificate)
Remarks:
Behörde für Gesundheit und Verbraucherschutz, Hamburg
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
other: CD / Crl: CD(SD)
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Females nulliparous and non-pregnant: yes
- Age at study initiation: Approx. 8 weeks
- Weight at study initiation: 171 - 193 g
- Fasting period before study: yes, 16 hours prior administration
- Housing: During the 14-day observation period the animals were kept in groups of 3 animals in MAKROLON cages (type III plus)
- Diet: Commercial diet, ssniff® R/M-H V1534; ad libitum
- Water: drinking water provided ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 15
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12 hours each
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 200 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw
- Justification for choice of vehicle: Corn oil was chosen as vehicle as it is known not to produce toxic effects.

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

CLASS METHOD
- Rationale for the selection of the starting dose: Three animals of one sex (preferably females) were first treated at 2000 mg/kg bw. If two to three animals died, testing at 300 mg/kg bw would be performed. If no to one animal dies, the test item was retested with 2000 mg/kg bw, using three animals of the same sex. If, in this second step, two to three animals died, testing at 300 mg/kg bw was performed. If, in this second step, no to one animal dies, no further testing was necessary.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
6 female rats
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Observations were performed before and immediately, 5, 15, 30 and 60 min, as well as 3, 6 and 24 hours after administration. All animals were observed for a period of 14 days. Individual body weights were recorded before administration of the test item and thereafter in weekly intervals up to the end of the study
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs (changes of skin and fur, eyes and mucous membranes, respiratory and circulatory function, autonomic and central nervous system and somatomotor activity as well as behaviour pattern)
Statistics:
No statistical analysis could be performed (the method used is not intended to allow a calculation of a precise LD50 value).
Key result
Sex:
female
Dose descriptor:
LD50 cut-off
Effect level:
5 000 mg/kg bw
Based on:
test mat.
Mortality:
No animal died prematurely.
Clinical signs:
No signs of toxicity were observed in any of the treated female rats.
Body weight:
All animals gained the expected body weight.
Gross pathology:
No pathological changes were observed at necropsy.
Interpretation of results:
other: CLP/EU GHS criteria are not met, no classification is required according to Regulations (EC) No 1272/2008.
Conclusions:
In an acute oral toxicity study conducted according to OECD 423, no mortality was observed in all treated female rats and the LD50 cut-off was established at 5000 mg/kg bw.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
18.05. - 4.07.2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Entidad Nacional de Acreditación
Test type:
acute toxic class method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS Spain S.L., Crta. Sant Miquel del Fai, km 3, 08182 Sant Feliú de Codines, Barcelona – Spain
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 8-9 weeks
- Weight at study initiation: Group A: males: mean of 224.5 g; females: mean of 193.9 g; Group B: males: mean of 282.9 g; females: mean of 169.0 g
- Fasting period before study: no but animals were deprived during exposure
- Housing: 4/cage (before distribution) and 3/cage (after distribution)
- Diet: Global diet provided ad libitum
- Water: Tap water provided ad libitum
- Acclimation period: 8 days (Group A) or 13 days (Group B) prior exposure

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.2 - 24.6
- Humidity (%): 25 - 63
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12h light : 12h dark
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Mass median aerodynamic diameter (MMAD):
>= 1 - <= 4 µm
Geometric standard deviation (GSD):
>= 1.5 - <= 3
Remark on MMAD/GSD:
Mass median aerodynamic diameter (MMAD) and the geometric standard deviation (GSD) were determined at the target concentration and calculated on the basis of the results from the cascade impactor, using Microsoft Excel® software (Microsoft Corporation, USA). The target ranges were 1 to 4 μm for the MMAD and 1.5 to 3 for the GSD. A respirable aerosol (MMAD in the range of 1-4 μm) could be achieved at 3.5 mg/L air. Therefore, starting dose was set at 3.5 mg/L although, as indicated above, the mean actual dose reached during the present study was 3.81 mg/L.
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Exposure chambers type EC-FPC-232
- Exposure chamber volume: approximately 3 L
- Method of holding animals in test chamber: restraint tubes which were positioned radially around the exposure chamber
- Source and rate of air: The flow of air at each tube was approximately 1.1 L/min, which is sufficient to minimize rebreathing of the test aerosol as it is more than twice the respiratory minute volume of rats.
- System of generating particulates/aerosols: A dust aerosol was generated from the desiccated and sieved test item using a Dust Generator SAG 410 (TOPAS GmbH, Germany)
- Method of particle size determination: Mass median aerodynamic diameter (MMAD) and the geometric standard deviation (GSD)
- Treatment of exhaust air: not reported
- Temperature, humidity, pressure in air chamber: temperature = 19 - 25 ºC; humidity = 30 - 70%;

TEST ATMOSPHERE
- Brief description of analytical method used: a stable aerosol of approximately 3.5 mg/L could be achieved with the TOPAS SAG 410 aerosol generator
- Samples taken from breathing zone: yes; test aerosol samples were collected onto a Whatman filter (grade F319.04) using a filter sampling device. The duration of sampling was 5 minutes.

Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
ca. 4 h
Concentrations:
3.81 and 1.02 mg/L
(3.81 mg/L was the highest stable aerosol concentration achievable that could be maintained at least for 4 hours.)
No. of animals per sex per dose:
3 animals/sex/group
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical observations in response to treatment were performed on all animals hourly during exposure (only grossly abnormal signs), immediately and 1 h after exposure, and once daily thereafter until the end of the observation period. Any visible clinical signs and discomfort were recorded. All animals were weighed once during the acclimatization period, on the day of treatment just before starting exposure, 24 h, 72 h and one week thereafter and also before sacrifice and gross necropsy conducted two weeks after exposure. Nevertheless, animals from Group A exposed to 3.81 mg/L were also weighed 4, 5 and 6 days after exposure for animal welfare reasons due to the mortality recorded and the severe clinical signs present.
- Necropsy of survivors performed: yes; the gross necropsy consisted of the examination of the abdominal and thoracic cavities and contents. Special attention was paid to any change in the respiratory tract. Any organs with gross lesions were collected and preserved in fixation medium (neutral-buffered 4% formaldehyde) for histological evaluation if considered relevant.
- Other examinations performed: Any clinical signs, discomfort and mortality were recorded in accordance with the humane endpoints guidance document of the OECD.
Statistics:
No statistical analysis was required.
Key result
Sex:
male/female
Dose descriptor:
LC50 cut-off
Effect level:
3.81 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Remarks on result:
other: 3.81 mg/L was the highest technically achievable concentration
Mortality:
In the high dose group (A, 3.81 mg/L), two male animals died shortly before finishing exposure and shortly thereafter, respectively, and the third one had to be sacrificed on study day 7 for animal welfare reasons due to the presence of severe labored breathing and continuous body weight loss (35% in total). All 3 females from Group A exposed to the dose of 3.81 mg/L survived the 14-day observation period after exposure.
No mortality was recorded in the low dose group (B, 1.02 mg/L).
Clinical signs:
other: The main clinical signs observed in both groups after finishing exposure were chromodacryorrhea, chromorrhinorrhea, soiled coat and piloerection. All these signs were transient and most of them were not present 24 hours after exposure. Breathing difficult
Body weight:
In all surviving animals, body weight decreased 24 hours after exposure. Afterwards, body weight tended to gradually increase in most animals until the end of the observation period, although in a number of cases slight decreases were also observed until 72-96 hours after exposure (two females from the high dose group and one female from the low dose group).
Gross pathology:
The necropsy conducted in both prematurely dead male animals from the high dose group, revealed a laryngeal obstruction by a white matter compatible with test item rests, whereas in the third male from this group euthanized for humane reasons on study day 7 a nodule of unknown origin was found on the outer tracheal wall. Red spots in the mandibular lymph nodes from both early decedent males and in the lungs from one of them were also observed.

No macroscopical findings were observed either in females from the high dose group or in animals exposed to the low dose, both sexes.
Interpretation of results:
other: CLP/EU GHS Category 4 (H302) according to Regulation (EC) No 1272/2008
Conclusions:
In an acute inhalation toxicity study conducted according to OECD 436, all male rats in the 3.81 mg/L dose group died. 3.81 mg/L was the highest technically achievable concentration. No mortality was observed in all female rats of the 3.81 mg/L dose group and all animals in the 1.02 mg/L dose group. Thus, the LC50-cut-off was found to be 3.81 mg/L.
Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017-01-17 to 2017-05-09
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Version / remarks:
February 24, 1987
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Version / remarks:
May 30, 2008
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Behörde für Gesundheit und Verbraucherschutz, Hamburg
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: CD / Crl: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Females nulliparous and non-pregnant: not reported
- Age at study initiation: Males: approx. 8 weeks; Females: approx. 9 weeks
- Weight at study initiation: Males: 247 - 256 g; Females: 229 - 251 g
- Housing: Individually housed in MAKROLON cages (type III plus)
- Diet: Commercial diet, ssniff® R/M-H V1534; not reported whether diet was provided ad libitum
- Water: drinking water provided ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 15
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12 hours each
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Area of exposure: shaved intact dorsal skin (5 cm x 6 cm, approx.)
- % coverage: 1/10 of body surface
- Type of wrap if used: layers of gauze covered with a plastic sheet and secured with adhesive plaster strips

REMOVAL OF TEST SUBSTANCE
- Washing: no
- Time after start of exposure: 24 h

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 3000 mg/kg bw for a dose of 2000 mg/kg bw
- For solids, paste formed: yes
Duration of exposure:
24 hours
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5 males and 5 females
Control animals:
not required
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Observations were performed before and immediately, 5, 15, 30 and 60 min, as well as 3, 6 and 24 hours after administration. All animals were observed for a period of 14 days. Individual body weights were recorded before administration of the test item and thereafter in weekly intervals up to the end of the study
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs (changes of skin and fur, eyes and mucous membranes, respiratory and circulatory function, autonomic and central nervous system and somatomotor activity as well as behaviour pattern)
Preliminary study:
No signs of toxicity were observed in animals administered 500, 1000 of 2000 mg Dimelamine pyrophosphate/kg bw.
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality
Clinical signs:
No signs of toxicity were noted. No skin reactions were observed at the application site.
Body weight:
All animals gained the expected body weight.
Gross pathology:
No signs of abnormalities were noted at necropsy.
Interpretation of results:
other: CLP/EU GHS criteria are not met, no classification is required according to Regulations (EC) No 1272/2008.
Conclusions:
In an acute dermal toxicity study conducted according to OECD 402, no mortality was observed in male and female rats and the LD50 was established at > 2000 mg/kg bw.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Additional information

Acute Oral

In an acute oral toxicity study (performed according to OECD 423 and GLP compliant), dimelamine pyrophosphate (CAS 13518-93-9) was examined for acute toxicity after a single oral administration of 2000 mg/kg bw to rats (LPT, 2017). Under the present test conditions, a single oral administration of 2000 mg Dimelamine pyrophosphate/kg b.w. to female rats did not reveal any signs of toxicity. All animals gained the expected body weight. No animal died prematurely. No pathological changes were observed at necropsy. The LD50 cut-off value was established at 5000 mg/kg bw.

Acute Inhalation

In an acute inhalation study (performed according to OECD 436 and GLP compliant), a group of three male and three female rats was exposed by nose-only, flow-past inhalation to an dust aerosol of dimelamine pyrophosphate (CAS 13518-93-9) at a mean concentration of 3.81 mg/L air during 4 hours (Vivotecnia, 2017). This concentration was found to be the highest technically achievable. However, due to the mortality found with this concentration, a second group of 3 male and 3 female animals was exposed at a mean concentration of 1.02 mg/L for 4 hours. As no animal exposed to the dose of 1.02 mg/L died with during the observation period, no additional doses were required.

In the high dose group (3.81 mg/L), two male animals died shortly before finishing exposure and shortly thereafter, respectively, and the third one had to be sacrificed on study day 7 for animal welfare reasons due to the presence of severe labored breathing and continuous body weight loss (35% in total). No mortality was recorded in the high dose group females and the low dose group (1.02 mg/L).

The main clinical signs observed in both groups after finishing exposure were chromodacryorrhea, chromorrhinorrhea, soiled coat and piloerection. All these signs were transient and most of them were not present 24 hours after exposure. Breathing difficulty was also present until sacrifice in the male animal which had to be euthanized and in the three females from the high dose group until study days 5 (two animals) and 8 (one animal). No clinical signs were observed in the animals from the low dose group, both sexes, from the day after exposure until the end of the observation period.

In all surviving animals, body weight decreased 24 hours after exposure. Afterwards, body weight gradually increased in most animals until the end of the observation period, although in a number of cases slight decreases were also observed until 72-96 hours after exposure (two females from the high dose group and one female from the low dose group).

The necropsy conducted in both prematurely dead male animals from the high dose group, revealed a laryngeal obstruction by a white matter compatible with test item rests, whereas in the third male from this group euthanized for humane reasons on study day 7 a nodule of unknown origin was found on the outer tracheal wall. Red spots in the mandibular lymph nodes from both early decedent males and in the lungs from one of them were also observed. No necropsy macroscopical findings were observed either in females from the high dose group or in animals exposed to the low dose, both sexes.

Based on these results, the LC50 cut-off value was established at 3.81 mg/L which is the highest technically achievable dose.

Acute Dermal

In an acute dermal toxicity study (performed according to OECD 402 and GLP compliant), dimelamine pyrophosphate (CAS 13518-93-9) was examined for acute toxicity after a single dermal administration of 2000 mg/kg bw to rats (LTP, 2017). Under the present test conditions, a single dermal administration of 2000 mg dimelamine pyrophosphate/kg b.w. did not reveal any signs of toxicity nor skin reactions at the application site. All animals gained the expected body weight. No animal died prematurely. The LD50 value was established at > 2000 mg/kg bw.

Justification for classification or non-classification

The available data on acute oral and dermal toxicity of dimelamine pyrophosphate do not meet the criteria for classification according to Regulation (EC) No 1272/2008, and are therefore conclusive but not sufficient for classification.

The available data on acute inhalation toxicity of dimelamine pyrophosphate meets the criteria for classification according to Regulation (EC) No 1272/2008, and is therefore classifiedy as Category 4 (H332).