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

Endpoint summary

Description of key information

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

Dermal: LD50 (rat, females)>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:

2021-04-22 to 2021-07-08

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

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

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.

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

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.

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.

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

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

5 000 mg/kg bw

Quality of whole database:

Guideline Study under GLP; high quality

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:

experimental study

Adequacy of study:

key study

Study period:

17 February 2022 to 11 March 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

OECD Guideline for Testing of Chemicals No. 402 (Section 4: Health Effects) “Acute Dermal Toxicity: Fixed Dose Procedure” adopted on 09 October 2017

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and batch No.of test material: Clariant and S148866
- Expiration date of the batch: 30.09.2022

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: No
- Final preparation of a solid: applied as such

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: In-house bred
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 11 weeks
- Weight at study initiation: 200.45 g to 201.69 g
- Fasting period before study: No
- Housing: polysulphonate cage (size: L 430 x B 280 x H 210 mm)
- Diet (e.g. ad libitum): Yes
- Water (e.g. ad libitum): Yes
- Acclimation period: 18 Feb 2022 to 24 Feb 2022

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.0°C to 22.8°C
- Humidity (%): 44% to 66%
- Air changes (per hr): 12 to 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light and 12 hours dark cycle

IN-LIFE DATES: From: 18 Feb 2022 To: 11 Mar 2022

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: approximately 6 cm × 10 cm
- % coverage: 10% of the total body surface
- Type of wrap if used: crepe bandage

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes
- Time after start of exposure: 24 Hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 420.3 mg for range-finding study; 439.3 and 440.3 mg for main study
- Constant volume or concentration used: yes

Duration of exposure:

24 hours

Doses:

2000 mg/kg body weight

No. of animals per sex per dose:

Range-finding study: 1 animal
Main study: 2 animals

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: observation once daily and weekly weighing
- Necropsy of survivors performed: yes
- Other examinations performed: clinical sign and body weight

Statistics:

No

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

not determinable due to absence of adverse toxic effects

Mortality:

No mortality

Clinical signs:

other: No treatment related clinical signs of toxicity and mortality were observed in both range finding study and main study animals

Gross pathology:

No treatment related gross pathological changes were observed in any of the animals in both range finding study and main study

 

TABLE 1.     INDIVIDUAL ANIMAL CLINICAL SIGNS OF TOXICITY ANDMORTALITY RECORD

Phase of the Experiment

Dose (mg/kg body weight)

Animal No.

Sex

Time of Application

Clinical Signs of Toxicity and Mortality on Day 1

Clinical Signs of Toxicity and Mortality on days

20-30 mins

1 hr (±10 mins)

2 hrs (±10 mins)

4 hrs (±10 mins)

6 hrs (±10 mins)

2

3

4

5

6

7

8

9

10

11

12

13

14

15

Range Finding Study

2000

Rg6745

F

11:44

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Main Study

2000

Rg6746

F

11:45

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rg6747

F

11:46

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

    N: Normal; F: Female; mins: minutes; hr/hrs: hour/hours

 

 TABLE 2.  INDIVIDUAL ANIMAL SKIN REACTION SCORE RECORDD

Phase of the Experiment	Dose (mg/kg body weight)	Sex	Animal No.	Skin Reaction Observations	Skin Reaction Score on Days
					3                (24 hour)	4                  (48 hour)	5             (72 hour)
Range Finding Study	2000	F	Rg6745	ER	0	0	0
				ED	0	0	0
Main Study	2000	F	Rg6746	ER	0	0	0
				ED	0	0	0
		F	Rg6747	ER	0	0	0
				ED	0	0	0

      F: Female; ER: Erythema; ED: Edema; 0: No erythema and no edem

 

 

 

TABLE 3.INDIVIDUAL ANIMAL BODY WEIGHT (g) AND PERCENT CHANGE IN BODY WEIGHT WITH RESPECT TO DAY 1

Phase of the Experiment	Dose (mg/kg body weight)	Animal No.	Sex	Quantity of test item applied (mg)	Body Weight (g) on Days				Percent Change in Body Weight with Respect to Day
					1	8	15		1 to 8	1 to 15
Range Finding Study	2000	Rg6745	F	420.3	210.15	228.61	245.13		8.78	16.65
Main Study	2000	Rg6746	F	439.3	219.65	236.13	252.73		7.50	15.06
		Rg6747	F	440.3	220.16	238.21	256.41		8.20	16.47
				Mean	219.91	237.17	254.57		7.85	15.76
				±SD	0.36	1.47	2.60		0.49	0.99
				n	2	2	2		2	2

     F: Female; SD: Standard Deviation; n: Number of animals

 

 

Quantity of Test Item Applied (mg)	=	Dose / 1000 X Body weight (g)
	=	2000 / 1000 X 210.15
	=	420.3 mg

 

 

 

 

TABLE 4.    INDIVIDUAL ANIMALGROSS PATHOLOGY FINDINGS

Phase of the Experiment	Dose (mg/kg body weight)	Animal No.	Sex	Fate	Gross Pathology Findings
					External	Internal
Range finding Study	2000	Rg6745	F	TS	NAD	NAD
Main Study	2000	Rg6746	F	TS	NAD	NAD
		Rg6747	F	TS	NAD	NAD

 NAD: No Abnormality Detected; F: Female;TS: Terminal Sacrifice

Interpretation of results:

GHS criteria not met

Conclusions:

Under the experimental conditions employed and based on the above results, it is concluded that the acute dermal median lethal dose (LD50) of test item, Cyclic Glucamide C12-C14 in Sprague Dawley rats is >2000 mg/kg body weight.

Executive summary:

 

The test item Cyclic Glucamide C12-C14 was evaluated for Acute Dermal Toxicity study in Sprague Dawley Rats.

The study was performed in two phases i.e. range finding study and main study. Range finding study was performed with one female rat and main study was performed with two female rats. On the day before the application of the test item, fur on the dorso-lateral area of the trunk of the animals was removed by clipping closely with an electric hair clipper and care was taken to avoid abrading the skin.

The required quantity of the test item was applied as uniform film over an area of approximately 10% of the total body surface. The test item was held on to the applied surface by covering with cotton gauze dressing and wrapped with non-irritating adhesive tape and finally the application site was wrapped using semi-occlusive crepe bandage. The contact period of test item was 24 hours. At the end of the contact period, the residual test item was washed using distilled water and dried with absorbent cotton.

The animals were dosed in a stepwise procedure with one female rat at a time in range finding study. Since the LD₅₀of structure analogues is >2000 mg/kg body weight based on information provided by sponsor and as per the material safety data sheet provided by sponsor the LD₅₀of test item is >2000 mg/kg body weight, a starting dose of 2000 mg/kg body weight was selected from the fixed dose levels of 50, 200, 1000 and 2000 mg/kg body weight. No clinical signs and mortalities were observed at the dose level of 2000 mg/kg body weight in range finding study. Hence, during main study, two animals were administered with the same dose level of 2000 mg/kg body weight. No clinical signs and mortalities were observed at the dose level of                 2000 mg/kg body weight. Hence, no further testing was carried out.

All the animals were observed for clinical signs of toxicity and mortality at               20 to 30 min, 1 hr (±10 mins), 2 hrs (±10 mins), 4 hrs (±10 mins) and 6 hrs                  (±10 mins) post dosing on day 1 and thereafter once daily for clinical signs of toxicity and twice daily for mortality during the 14 days observation period. Skin reactions were scored at 24, 48 and 72 hours after patch removal using draize scoring system. The body weight was recorded at receipt, on day 1 before test item application and on days 8 and 15. At the end of observation period, all the animals were humanely sacrificed by carbon dioxide asphyxiation and subjected to necropsy and detailed gross pathological examination.

No mortality and clinical signs were noted.

No skin reactions were noted at 24, 48 and 72 hours after patch removal.

No treatment related changes in body weight and percent change in body weight with respect to day 1 were noted. Normal increase in body weights were noted during the observation period. 

No treatment related gross pathological changes were noted at 2000 mg/kg body weight (range finding study and main study) during necropsy

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

> 2 000 mg/kg bw

Quality of whole database:

Guideline Study under GLP; high quality

Justification for classification or non-classification

Based on the results from available guideline studies which revealed no adverse effects, the registration substance is not subject to labelling and classification requirements.