Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
April-June 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
Name: RCX 15-116
CAS No.: 15305-07-4
Batch No.: 15047AE2
Purity: > 99%
Chemical Name: Aluminium-N-nitrosophenylhydroxylamine (NPAL)
Aggregate State at RT: solid
Colour: whitish
Storage Conditions: room temperature, protected from light
Stability: Instable after repeated contact to air and / or light, undergoes hydrolysis
Expiry Date: 30 Nov 2016
Safety Precautions: The routine hygienic procedures were sufficient to assure personnel health and safety.

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
Species/strain: WISTAR rats Crl: WI(Han)
Source: Charles River, 97633 Sulzfeld, Germany
Sex: female (non-pregnant and nulliparous)
Number of animals: 3 per step
Age at the
beginning of the study: 9-12 weeks old
Body weight on the
day of administration: step 1: animals no.1 – 3: 167 - 178 g;
step 2: animals no.4 – 6: 177 - 185 g;
step 3: animals no.7 – 9: 152 - 170 g


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
- 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 in groups in IVC cages, type III H, polysulphone cages on Altromin saw fibre bedding
- 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

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Prior to the administration food was withheld from the test animals for 16 to 19 hours (access to water was permitted). Following the period of fasting the animals were weighed and the test item was administered. Food was provided again approximately 4 hours post dosing.

The test item was administered at a single dose by gavage using a feeding tube.The test item was administered at a dose volume of 10 mL/kg body weight.

The starting dose was selected to be 2000 mg/kg body weight. Compound related mortality was recorded for all animals of step 1. Based on these results and according to the acute toxic class method regime, a second step was performed at a dose of 300 mg/kg body weight. No compound related mortality was recorded for any animal of step 2. Based on these results and according to the acute toxic class method regime, a third step was performed at a dose of 300 mg/kg body weight. No compound-related mortality was recorded for any animal of step 3. Based on these results and according to the acute toxic class method regime no further testing was required.
Doses:
2000 and 300 mg/kg bw
No. of animals per sex per dose:
3
Control animals:
no
Details on study design:
The surviving animals were observed for 14 days after dosing for general clinical signs, morbidity and mortality. The animals were weighed on day 1 (prior to the administration) and on days 8 and 15.A 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. Particular attention was directed to observations of tremor, convulsions, salivation, diarrhoea, lethargy, sleep and

The animals which died spontaneously during the observation period were necropsied as soon as they was found. At the end of the observation period the surviving animals were sacrificed with an overdosage of pentobarbital injected intraperitoneally at a dosage of 250-400 mg/kg bw. All animals were subjected to gross necropsy and examined macroscopically for gross pathological changes. In the absence of gross pathological changes no tissues were preserved for a possible histopathological evaluation.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50 cut-off
Effect level:
500 mg/kg bw
Based on:
test mat.
Mortality:
All animals treated with the test item at a dose of 2000 mg/kg died spontaneously on the day of treatment. All remaining animals survived until the end of the study showing signs of toxicity.
Clinical signs:
The most relevant clinical findings in the animals treated with the test item at a dose of 2000 mg/kg bw were reduced spontaneous activity, prone position, slow movements, ataxia, hunched posture, cyanosis, eyes half closed, eyes closed and abnormal breathing. The most relevant clinical findings in the animals treated with the test item at a dose of 300 mg/kg bw were reduced spontaneous activity, ataxia, cyanosis and eyes half closed. All symptoms recovered within 1 day post-dose.
Body weight:
Throughout the 14-day observation period, the weight gain of the surviving animals was within the normal range of variation for this strain.
Gross pathology:
Macroscopic findings of surviving animals: at necropsy, no treatment-related macroscopic findings were observed in any animal of any step.
Macroscopic findings of animals not having survived until the end of the observation period:necropsy revealed a bluish discoloured lung.

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
Under the conditions of the present study, a single oral application of the test item RCX 15-116 to rats at a dose of 2000 mg/kg body weight was associated with signs of toxicity and mortality. A single oral application of the test item RCX 15-116 to rats at a dose of 300 mg/kg body weight was associated with signs of toxicity but no mortality. The median lethal dose of RCX 15-116 after a single oral administration to female rats, observed over a period of 14 days is: LD50cut-off (rat): 500 mg/ kg bw.
Executive summary:

The acute oral toxicity study was conducted according to OECD Guideline 423 (Acute Toxic Class Method) and under GLP. One groups of three female WISTAR Crl: WI(Han) rats was treated with the test item by oral gavage administration at a dosage of 2000 mg/kg body weight. The test item was suspended with the vehicle corn oil at a concentration of 0.2 g/mL and administered at a dose volume of 10 mL/kg. Two groups, each of three female WISTAR Crl: WI(Han) rats, were treated with the test item by oral gavage administration at a dosage of 300 mg/kg body weight. The test item was suspended with the vehicle corn oil at a concentration of 0.03 g/mL and administered at a dose volume of 10 mL/kg. The most relevant clinical findings in the animals treated with the test item at a dose of 2000 mg/kg bw were reduced spontaneous activity, prone position, slow movements, ataxia, hunched posture, cyanosis, eyes half closed, eyes closed and abnormal breathing. The most relevant clinical findings in the animals treated with the test item at a dose of 300 mg/kg bw were reduced spontaneous activity, ataxia, cyanosis and eyes half closed. All symptoms recovered within 1 day post-dose. Throughout the 14-day observation period, the weight gain of the surviving animals was within the normal range of variation for this strain. The LD50 (cut-off) was determined to be 500 mg/kg bw.