cyclic C12/14-Glucamide;cyclic N-methyl-N-dodecanoyl/tetradecanoyl-glucamine;anhydro-N-dodecanoyl/tetradecanoyl-N-methylglucamine;Amides, C12-C14, N-(1-deoxy-D-glucitol-1-yl)-N-methyl, dehydrated

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2021-04-22 to 2021-07-08

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

D-Glucitol, anhydro-1-deoxy-1-(methylamino)-, N-[C12-14(even numbered) acyl] derivs.

Molecular formula:

C19H37NO5 C21H41NO5

IUPAC Name:

D-Glucitol, anhydro-1-deoxy-1-(methylamino)-, N-[C12-14(even numbered) acyl] derivs.

Test material form:

semi-solid (amorphous): gel

Specific details on test material used for the study:

Name: cyclic Glucamide C12-C14
Product
(Common Name/Code): cyclic N-dodecanoyl/tetradecanoyl-N-methylglucamine / cyclic Glucamide C12-C14
Lot No.: S148866
Storage Conditions: room temperature
Physical State at Room
Temperature: slightly turbid solid mass
Color: brown
Density: 1.07 g/cm³
Molecular Weight: main compounds: 359,5 g/mol (C12-derivative); 387,5 g/mol (C14-derivative)
pH at at Room
Temperature: approx. 6-8 (not tested)
Date of Analysis: 30 September 2020
Expiry Date: 30 September 2022
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 or test system and environmental conditions:

Test System
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: 8 – 10 weeks
Body weight on the
day of administration:Step 1: 162 – 168 g;
Step 2: 178 – 186 g
The animals were derived from a controlled full-barrier maintained breeding system (SPF). According to the German Act on Animal Welfare [8] the animals were bred for experimental purposes.
This study was performed in an AAALAC-accredited laboratory. According to German animal protection law, the study type has been reviewed and accepted by local authorities. Furthermore, the study has been subjected to Ethical Review Process and was authorised by the Bavarian animal welfare administration.


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 Product Testing Munich GmbH
- Adequate acclimatisation period (at least five days) under laboratory conditions

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Aqua ad injectionem (sterile water, Deltamedica, lot no. 2005066, expiry date: 04/2023). This vehicle was chosen due to its non-toxic characteristics.

Details on oral exposure:

Preparation of the Animals

The animals were marked for individual identification by tail painting.
Prior to the administration a detailed clinical observation was made of all animals. Only healthy animals were used.
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 3 hours post dosing.

Administration

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.

Volume of Test Item Preparation Applied per Animal
Animal No. / Sex Dose (mg/kg bw) Body Weight on Day of Administration (g) Volume of Test Item Preparation in Vehicle
Administered per Animal (mL)
1 / Female 2000 162 1.6
2 / Female 168 1.7
3 / Female 165 1.7
4 / Female 2000 178 1.8
5 / Female 183 1.8
6 / Female 186 1.9

Doses:

The starting dose was selected to be 2000 mg/kg body weight. No compound-related mortality was recorded for any animal of step 1 or 2. Based on these results and according to the acute toxic class method regime no further testing was required.

No. of animals per sex per dose:

3 per step / 2 steps performed

Control animals:

no

Details on study design:

Observation Period
All animals were observed for 14 days after dosing for general clinical signs, morbidity and mortality.

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

Clinical Examination
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 coma.

Pathology
At the end of the observation period the animals were sacrificed with an overdosage of pentobarbital injected intraperitoneally at a dosage of 400-800 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.
 
Evaluation of Results
Results were interpreted according to OECD Guideline 423, Annex 2 and GHS.
Individual reactions of each animal were recorded at each time of observation.
Toxic response data were recorded by 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.

Results and discussion

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50 cut-off

Effect level:

>= 5 000 mg/kg bw

Based on:

test mat.

Mortality:

All animals survived until the end of the study showing slight signs of toxicity.

Clinical signs:

other: The most relevant clinical findings in the animals were reduced spontaneous activity, hunched posture, piloerection, prone position, half eyelid closure and eyes closed. All animals recovered within up to 3 days post-dose.

Gross pathology:

At necropsy, no macroscopic findings were observed in any animal of any step.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the present study, a single oral application of the test item Cyclic Glucamide C12-C14 to rats at a dose of 2000 mg/kg body weight was associated with signs of toxicity but no mortality.
The median lethal dose of Cyclic Glucamide C12-C14 after a single oral administration to female rats, observed over a period of 14 days is:
LD50 cut-off (rat): 5000 mg/ kg bw

Executive summary:

Summary Results

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 2000 mg/kg body weight.On the sponsors request and based on the outcome of the solubility test, the test item was dissolved in the vehicle aqua ad injectionem (sterile water) at a concentration of 0.2 g/mL and administered at a dose volume of 10 mL/kg.

All animals used in the study after their entrance at BSL were allowed to acclimatise to the laboratory conditions for at least 5 days. The animals were observed on delivery, on inclusion in the study and before administration for mortality/morbidity and other clinical signs. All animals were examined for clinical signs several times on the day of dosing and once daily until the end of the observation period. Their body weights were recorded on day 1 (prior to the administration) and on days 8 and 15.All animals were necropsied and examined macroscopically.

Table1:  Resultsper Step

Step		Sex / No.		Starting Dose (mg/kg bw)		Number of Animals		Number of Intercurrent Deaths
1		Female / 1 - 3		2000		3		0
2		Female / 4 - 6		2000		3		0

All animals survived until the end of the study showing slight signs of toxicity.

The most relevant clinical findings in the animals were reduced spontaneous activity, hunched posture, piloerection, prone position, half eyelid closure and eyes closed. All animals recovered within up to 3 days post-dose.

Throughout the 14-day observation period, the weight gain of the animals was within the normal range of variation for this strain.

At necropsy, no macroscopic findings were observed in any animal of any step.

LD₅₀cut-off (rat):                    5000 mg/kg bw

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

Vehicle:                                  aqua ad injectionem (sterile water)

Number of animals:                3 per step / 2 steps performed

Method:                                 OECD 423, EC 440/2008, Method B.1 tris, OPPTS 870.1100

 

 Conclusion

Under the conditions of the present study, a single oral application of the test item Cyclic Glucamide C12-C14 to rats at a dose of 2000 mg/kg body weight was associated with signs of toxicity but no mortality.

The median lethal dose of Cyclic Glucamide C12-C14 after a single oral administration to female rats, observed over a period of 14 days is:

LD₅₀cut-off (rat): 5000 mg/ kg bw