2-[[(2S)-2-(9H-fluoren-9-ylmethoxycarbonylamino)-3-(1H-imidazol-5-yl)propanoyl]amino]-2-methyl-propanoic acid Trifluoroacetic acid salt (1:1)

Endpoint summary

Description of key information

The oral LD50 for female rats was estimated to be 5000 mg.Kg-1 of body weight.

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:

25/03/2019 - 16/05/2019

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)

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

no

Specific details on test material used for the study:

Batch: 160004
Preparation date: 30/11/2016
Expiration date: 09/02/2020
Storage conditions: Refrigerated (2-8 °C)

Species:

rat

Strain:

Brown Norway

Remarks:

Rattus norvegicus

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: ANILAB - Animais de Laboratório Criação e Comércio Ltda
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 1st and 2nd treatmente group: 8 weeks; 3rd and 4th treatment group: 9 weeks
- Weight at study initiation: 1st treatment group: 205.88 to 222.75 g; 2nd treatment group: 191.53 to 205.96 g; 3rd treatment group: 177.94 to 216.02 g; 4th treatment group: 177.45 to 210.75 g
- Fasting period before study: The feed supplied to the animals was interrupted at the end of the day previous to the application of the test item.
- Housing: The animals were maintained in groups of 3 animals per treatment in polypropylene cage closed with a metallic grid, papered with shavings of Pinnus.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Min: 19.5ºC, Max: 23.4ºC
- Humidity (%): Min: 48%, Max: 70%
- Air changes (per hr): 10 to 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 6.6660 mg of test item resulting in a concentration of 1.1110 g.mL-1
- Amount of vehicle: 6 mL of purified water
- The administration was performed by gavage, using a suitable metal cannula attached to a syringe.
- Lot/batch no.: 160004
- Purity: ca. 100%

Doses:

300 mg/kg (1st and 2nd treatment group); 2000 mg/Kg bw (3rd and 4th treatment group);

No. of animals per sex per dose:

3 female rats/treatment group

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of weighing: Day 0, 7 and14
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other: yes

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

ca. 5 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: estimated on the basis of the flow chart in the OECD Guideline 423 Annex 2

Mortality:

No treatment-related deaths at the treatment doses.

Clinical signs:

other: No behavioral and clinical alterations observed during the experimental period at the treatment doses. Please refer to table 2 attached below.

Other findings:

Please refer to table 3 attached below.

Table 1 -Individual body weight and information about the test item administred.

Body Weight (g)
Treatment	Dose (mg.Kg^-1)	Animal #	Initial (Day 0)	Day 7	Day 14	Difference between final and initial weight	Administered quantity (mL)	Date and hour of application
1st	300	1	205.88	218.60	229.15	23.27	0.12	16/Apr/2019 09:15 a.m.
		2	222.75	251.76	261.00	38.25	0.12
		3	212.21	22.92	229.61	17.40	0.12
2nd	300	1	202.99	207.56	218.36	15.37	0.12	18/Apr/2019 09:42 a.m.
		2	205.96	212.08	224.76	18.80	0.12
		3	191.53	200.35	216.31	24.78	0.10
3rd	2000	1	191.11	196.63	198.99	7.88	0.68	23/Apr/2019 09:00 a.m.
		2	216.02	228.24	227.31	11.29	0.76
		3	177.94	190.56	195.84	17.90	0.62
4th	2000	1	210.75	217.77	219.06	8.31	0.84	25/Apr/2019 08:30 a.m.
		2	177.45	181.97	185.48	8.03	0.70
		3	189.79	200.62	2014.35	14.56	0.74

  

Table 2 -Behavioral and clinical alterations observed during the experimental period.

				observation day
Treatment	Step	Sex	Animal #	0				1	2	3	4	5	6	7	8	9	10	11	12	13	14
				0:30h*	10:30 a.m.	12:20 p.m.	02:00 p.m.
300	1	♀	1	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			2	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			3	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
				observation day
Treatment	Step	Sex	Animal #	0				1	2	3	4	5	6	7	8	9	10	11	12	13	14
				0:30h*	11:00 a.m.	12:50 p.m.	03:00 p.m.
300	2	♀	1	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			2	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			3	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
				observation day
Treatment	Step	Sex	Animal #	0				1	2	3	4	5	6	7	8	9	10	11	12	13	14
				0:30h*	10:30 a.m.	12:00 p.m.	02:14 p.m.
2000	3	♀	1	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			2	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			3	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
				observation day
Treatment	Step	Sex	Animal #	0				1	2	3	4	5	6	7	8	9	10	11	12	13	14
				0:30h*	10:30 a.m.	12:00 p.m.	02:14 p.m.
2000	4	♀	1	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			2	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
			3	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0

Legend:

0. none visual alterations observed; 1. Skin, pile and eyes alterations; 2. Mucous membranes alterations; 3. Respiratory system alteration; 4. System circulation alteration; 5. Nervous system alteration; 6. Behavior pattern alteration;

7. Convulsions; 8. Salivation; 9. Diarrhea; 10. Lethargy; 11. Tremble; 12. Death. *. After application of test item

  

Table 3 -Pathological findings in animals at doses of 300 and 2000 mg.kg^-1 of body weight.

			Macroscopic Alterations
Treatment	Dose (mg.Kg-1)	Animal #	Skin	Brain	Eyes	Lungs	Heart	Liver	Spleen	Urinary system	G.I.T	R.T	Carcass
1st	300	1	0	0	0	0	0	0	0	0	0	0	0
		2	0	0	0	0	0	0	0	0	0	0	0
		3	0	0	0	0	0	0	0	0	0	0	0
2nd	300	1	0	0	0	0	0	0	0	0	0	0	0
		2	0	0	0	0	0	0	0	0	0	0	0
		3	0	0	0	0	0	0	0	0	0	0	0
3rd	2000	1	0	0	0	0	0	0	0	0	0	0	0
		2	0	0	0	0	0	0	0	0	0	0	0
		3	0	0	0	0	0	0	0	0	0	0	0
4th	2000	1	0	0	0	0	0	0	0	0	0	0	0
		2	0	0	0	0	0	0	0	0	0	0	0
		3	0	0	0	0	0	0	0	0	0	0	0

Legend:

G.I.T- Gastrointestinal tract

R.T - Reproductive tract

0 - Not observed alteration

A - Observed alteration

 

Interpretation of results:

Category 5 based on GHS criteria

Conclusions:

Under the test conditions, the test item FMOC-HIS-AIB-OH TFA LS;N-[(9HFLUOREN-9-YLMETHOXY) CARBONYL]-LHISTIDYL-2-METHYLALANINE ANDTRIFLUOROACETIC ACID (1:1); N.CAS:1446013-08-6- BATCH: 160004, when administered by oral route in female rats, did not cause deaths, for every step taken at the dose levels of 300 and 2000 mg.Kg-1 of body weight. In clinical examinations, the tested animals did not show systemic signs of toxicity. In macroscopic evaluations no alterations were observed during the necropsies. Based on the flow chart with the starting dose of 300 mg.Kg-1 of body weight, the test item was classified as category 5 (unclassified), according to the GHS (Globally Harmonized Classification System for Chemical Substances and Mixtures). The oral LD50 value of test item FMOC-HIS-AIB-OH TFA LS;N-[(9HFLUOREN-9-YLMETHOXY) CARBONYL]-LHISTIDYL-2-METHYLALANINE ANDTRIFLUOROACETIC ACID (1:1); N.CAS:1446013-08-6- BATCH: 160004, for female rats, was estimated to be 5000 mg.Kg-1 of body weight.

Executive summary:

This acute oral toxicity study in rats (Rattus norvegicus) was carried out in order to evaluate the possible toxic effects of the test item FMOC-HIS-AIB-OH TFA LS;N-[(9HFLUOREN-9-YLMETHOXY) CARBONYL]-LHISTIDYL-2-METHYLALANINE AND TRIFLUOROACETIC ACID (1:1); N.CAS:1446013-08-6- BATCH: 160004 administered by the oral route. Twelve females, divided in groups of three animals, were tested in four steps at the dose levels of 300 and 2000 mg.Kg-1 of body weight. The test item was applied diluted using purified water as vehicle. The volume administered to each animal was calculated according to the body weight determined on the day of treatment. After dosing by gavage, the animals were observed during 14 days to evaluate deaths, and behavioral and clinical alterations. The test item administered by oral route for female rats did not cause treatment-related deaths in any of the steps at the dose levels of 300 and 2000 mg.Kg-1 of body weight. At the clinical examinations, toxicity signs were not observed. The animals were euthanized in the end of the observation period and were submitted to necropsies, where they did not present macroscopic alterations or acute toxic effects caused by test item. Based on the flow chart with the starting dose of 300 mg.Kg-1 body weight, the test item was classified as category 5, according to the GHS (Globally Harmonized Classification System for Chemical Substances and Mixtures). The acute oral LD50 value of the test item FMOC-HIS-AIB-OH TFA LS;N-[(9HFLUOREN-9-YLMETHOXY) CARBONYL]-LHISTIDYL-2-METHYLALANINE AND TRIFLUOROACETIC ACID (1:1); N.CAS:1446013-08-6- BATCH: 160004 was estimated 5000 mg.Kg-1 of body weight for female rats.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

5 000 mg/kg bw