Pyrimidine-2,4,6-triyltriamine

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May - July 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1980

Report date:

1980

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Version / remarks:

Equivalent to Version dated May 1981

Deviations:

yes

Remarks:

- Acclimatization period was minimum 4 days, instead of at least 5 days as recommended by guideline - Volume should not exceed 10 ml/kg (OECD), in the study 12.5 ml/kg were used as max - No constant volume was used (concentrations were not adjusted)

Principles of method if other than guideline:

The study was performed prior to the implementation of the OECD test guideline 401 in May 1981. However, procedures are very similar to the later enforced guideline and was thus concluded to be equivalent to the test guideline with acceptable restrictions.

GLP compliance:

no

Remarks:

pre-GLP

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Pyrimidine-2,4,6-triyltriamine

EC Number:

213-720-7

EC Name:

Pyrimidine-2,4,6-triyltriamine

Cas Number:

1004-38-2

Molecular formula:

C4H7N5

IUPAC Name:

2,4,6-Triaminopyrimidine

Test material form:

solid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source: Lonza AG
- Purity: 98.7%
- Melting point: 254-256.5°C

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Final dilution of a dissolved solid: 40% w/v

FORM AS APPLIED IN THE TEST: suspension

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River UK Limited, Margate, Kent, England
- Females nulliparous and non-pregnant: yes (virgin)
- Age at study initiation: approx. 7 weeks
- Weight at study initiation: 170-180 g
- Fasting period before study: Approx. 18 +/- 2 hours prior to and 4 hours after dosing
- Housing: groups
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: minimum 4 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 +/- 2
- Humidity (%): 47 (median)
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To:

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 40% w/v suspension
- Amount of vehicle (if gavage): depending on dose. (max volume applied 12.5 ml/kg)

MAXIMUM DOSE VOLUME APPLIED: 12.5 ml/kg

DOSAGE PREPARATION: The test sample was prepared freshly on the day of dosing

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose:

Doses:

1000, 2500, and 5000 mg/kg bw applied as 40% w/v suspension

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

PRELIMIARY STUDY:

- establishing a dosing regimen
- two males and two females per dose level
- dosage levels: 1000, 2500, and 5000 mg/kg bw
- Duration of observation period following administration: 15 or 21 days

MAIN STUDY:

- Concentration of test substance in the vehicle was the same for all dosage levels
- Dosed volume was varied to achieve necessary dosage level
- Duration of observation period following administration: 15 days
- Frequency of observations and weighing: Observed soon after dosing, then hourly on day 1. Subsequnetly, animals were observed once in the morning and once at the end of the experimental day. Clinical signs were observed at each observation.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight

Statistics:

Acute oral median LD50 was calculated using the method of Finney (1971) Probit Analysis (3rd Edition) Cambridge University Press.

Results and discussion

Preliminary study:

Results of the preliminary study indicated that the LD50 was in the region of 2500 to 5000 mg/kg bw.

Effect levels

Effect levels 1

Sex:

female

Dose descriptor:

LD50

Effect level:

2 050 mg/kg bw

Based on:

test mat.

95% CL:

> 733 - < 3 241

Effect levels 2

Sex:

male

Dose descriptor:

LD50

Effect level:

2 500 mg/kg bw

Based on:

test mat.

Remarks on result:

other: could not be calculated from the mortality pattern, but appeared to be in the region of 2500 mg/kg bw.

Mortality:

Death occurred amongst rats at 1000 mg/kg bw and above from within one and ten days of dosing (see table 1). At 3200 mg/kg bw, one male was killed in extremis on Day 7. Autopsy revealed amongst these animals congestion and hemorrhages in the lungs and pallor of the liver, spleen and kidneys.

Clinical signs:

other: In all treated rats, pilo-erection and abnormal body carriage (hunched posture) shortly after dosing was observed. Recovery of the survivors as judged by external appearance and behavior was apparently complete within 14 days of dosing with the exception

Gross pathology:

At 1000, 3200, and 5000 mg/kg bw, a brown or white creamy coloration of the gastric contents amongst rats. Gaseous distension of the stomach and a red coloration of the peritoneum in one male and one female, respectively, at 5000 mg/kg bw. Dark coloration of the liver in one female at 1000 mg/kg bw and enlarged, pale adrenals in two females at 3200 mg/Kg bw. A swollen gelatinous preputial gland and undersized seminal vesicles, small spleen with granular surface, congestion of the peritoneal blood vessel and hemorrhages in the lunggs in the male at 3200 mg/kg bw that was killed in extremis. Terminal autopsy findings were within normal limits.

Any other information on results incl. tables

Table 1: Mortality data for groups of rats - preliminary study

Dosage (mg/kg bw)	Mortality ratio			Time of death after dosing (days)
	Male	Female	Combined
1000	0/2	0/2	0/4	-
2500	1/2	1/2	2/4	<2 - <3
5000	1/2	1/2	2/4	<1 - <3

Table 2: Mortality ratio and group mean body weight (g) of rats - main study

Sex	Dosage (mg/kg bw)	Body weight (g) at			Mortality ratio	Time of death after dosing (days)
		Dosing	1 week	2 weeks
Male	1000	188	260	304	0/5	-
	2500	187	251	299	2/5	<4
	3200	192	148+	DIED	5/5	<2 – 6
	5000	174	DIED	-	5/5	<2 – 3
Female	1000	168	202	221	1/5	<1
	2500	175	153	220	2/5	<4
	3200	168	118+	159	4/5	<2 - <4
	5000	166	115+	DIED	5/5	<1 - <10

 

+ = One rat only

Table 3: Signs of reaction to treatment ratio of rats - main study

Signs	Signs of reaction ratio
	Dose (mg/kg bw)
	1000	2500	3200	5000
Pilo-erection	10/10	10/10	10/10	10/10
Abnormal body carriage (hunched posture)	10/10	10/10	10/10	10/10
Abnormal gait (waddling)	3/10	10/10	10/10	10/10
Lethargy	1/10	10/10	10/10	10/10
Decreased respiratory rate	0/10	2/10	10/10	10/10
Increased lacrimation (discolored red)	0/10	1/10	5/10	5/10
Ptosis	0/10	3/10	2/10	3/10
Abnormal gait (walking on toes)	0/10	3/10	2/10	1/10
Diarrhoea	0/10	3/10	0/10	1/10
Diuresis	0/10	2/10	0/10	1/10
Pallor of the extremities	0/10	0/10	5/10	3/10
Fine body tremors	0/10	0/10	3/10	3/10
Coarse body tremors	0/10	0/10	0/10	1/10
Ataxia	0/10	0/10	0/10	1/10
Gasping respiration	0/10	0/10	0/10	1/10
Increased salivation	0/10	0/10	0/10	1/10
Penis swollen and enlarged	0/10	0/10	1/10	0/10

Applicant's summary and conclusion

Interpretation of results:

Category 5 based on GHS criteria

Conclusions:

The acute median lethal oral dose (LD50) and its 95% confidence limits to rats of the test substance were calculated to be 2050 (733 to 3241) mg/kg bw, 2156 (1486 to 2735) mg/kg bw, and 2500 mg/kg bw for females only, males and females combined, and males only in the region of this value, respectively.

Executive summary:

An acute oral toxicity test was performed in order to investigate the toxic potential of the test substance. Ten (five male and five female) Wistar rats were dosed to 1000, 2500, 3200, and 5000 mg/kg bw of the test substance. The concentration of the test substance was 40% w/v in corn oil. Body weight was meaured at dosing and then weekly thereafter. At 2500 mg/kg bw, two out of five males died. At 3200 mg/kg bw and above, all males (5/5) died. At 1000 mg/kg bw, one out of five females died. Two out of five, four out of five, and all females died at 2500, 3200, and 5000 mg/kg, respectively. Clinical signs, auch as abnormal gait (waddling) and lethargy were observed at 1000 mg/kg bw (females only) and above (males and females). Other clinical signs included decreased respiratory rate, fine body tremors, ataxia, and diarrhoea, among others. The LD50 for males and females was 2500 mg/kg bw and 2050 mg/kg bw, respectively. LD50 were calculated according to mortality patterns, however, for females only because male LD50 could not be calculated from mortality patterns.