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Toxicological information

Acute Toxicity: oral

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Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From June 06, 1990 to Aug. 03, 1990
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1990
Report Date:
1990

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Version / remarks:
Adopted : 24 February 1987
Deviations:
yes
Remarks:
No certificate of analysis and details on test substance available.
Principles of method if other than guideline:
Acute oral toxicity or LD50 value was determined by oral administration of test substance to male and female rats. Subsequently, observations of effects such as clinical signs and mortality were made for 14 days. At the end of observation period, all surviving animals were necropsied.
GLP compliance:
yes (incl. certificate)
Remarks:
(according to OECD principles of GLP)
Test type:
standard acute method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material: Lehmannblausulfat; 2- amino-4-hydroxyethylaminoanisole sulfate
- Substance type: Pure active substance
- Physical state: Grey powder
- Stability under test conditions: Stable
- Storage condition of test material: Ambient, in dark

Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: not specified
- Expiration date of the lot/batch: not specified
- Purity test date: not specified

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At ambient temperature
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: 1.5% to 10% in aqua deion

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: not specified
- Preliminary purification step (if any): not specified
- Final dilution of a dissolved solid, stock liquid or gel: 1.5 to 10% in aquia deion
- Final preparation of a solid: not specified

Test animals

Species:
mouse
Strain:
other: BOR: NMRI, white mice
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Firma Winkelmann, Versuchstierzucht, Borchen
- Age at study initiation: Not reported
- Weight at study initiation: Males: 24.8 - 37.8 g; Females: 21.7 - 29.4 g
- Fasting period before study: Animals were fasted from 16 hours before until 3-4 hours after administration of the test substance.
- Housing: Collectively housed up to a maximum of 5 animals per cage (Makrolon type III).
- Diet: Ssniff- R Alleindiet pellets; ad libitum
- Water: Drinking water as for human consumption in Makrolon drinking bottles; ad libitum
- Acclimation period: At least for 7 days

ENVIRONMENTAL CONDITIONS
- Temperature: 21 ± 2°C
- Relative humidity: 50 - 85 % (measured with thermohygrometer twice daily)
- Air changes: Not reported
- Photoperiod: 12 hours dark / 12 hours light, artificial lighting (120 lux) from 7.00 a.m. - 7.00 p.m.

EXPERIMENT INITIATION DATE: June 26, 1990
EXPERIMENT COMPLETION DATE: Aug. 03, 1990

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: Aqua deion
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 1.25 - 10%

MAXIMUM DOSE VOLUME APPLIED:0.38 mL
ADMINISTRATION OF TEST SUBSTANCE: A single oral administration of the test substance was performed by gavage using a stomach tube

- Rationale for the selection of the starting dose: Based on the results of range finding study doses for main study were selected.
Doses:
125, 250, 500, 750 and 1000 mg/kg bw.
No. of animals per sex per dose:
5 animals/sex/dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical observations were conducted at 10 min and 1, 2, 6, 24 hours and thereafter once daily up to Day 14. Body weights were measured at Days 0, 7 and 14.
- Necropsy of survivors performed: Yes, surviving animals were sacrificed by cervical dislocation after 14 days and gross pathological examinations were subsequently performed.

Results and discussion

Preliminary study:
A preliminary range finding test with dose of 2000 mg/kg bw was conducted on 2 female mice. Both the treated mice died at 2000 mg/kg bw.
Effect levelsopen allclose all
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
327 mg/kg bw
Based on:
act. ingr.
Key result
Sex:
male
Dose descriptor:
LD50
Effect level:
333 mg/kg bw
Based on:
act. ingr.
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
351 mg/kg bw
Based on:
act. ingr.
Mortality:
The following mortalities were observed within 24-72 hours after dosing:
1) At 125 mg/kg: 1/5 males and 1/5 females
2) At 250 mg/kg: 2/5 males and 1/5 females
3) At 500 mg/kg : 4/5 males and 3/5 females
4) At 750 mg/kg: 3/5 males and 5/5 females
5) At 1000 mg/kg: 5/5 males and 4/5 females
Details of pre-terminal deaths were provided in the table 2 under ‘Any other information on results incl. tables’
Clinical signs:
Severe clinical symptoms related to CNS symptoms, coordination, reflexes and autonomic functions were observed with dose related intensity as follows:
1) At 125 mg/kg: Decreased activity and pilorection
2) At 250 mg/kg: Decreased activity, ventral position, squatting position, pilorection
3) At 500 mg/kg: Decreased activity, abnormal gait, ventral position, squatting position, decreased body tone, decreased plantary reflex, increased cyanosis, pilorection, increased respiratory rate
4) At 750 mg/kg: Decreased activity, squatting position, abnormal body posture, increased cyanosis, pilorection, increased respiratory rate
5) At 1000 mg/kg: Decreased activity, squatting position, abnormal body posture, righting reflex, increased cyanosis, pilorection, increased respiratory rate.
Body weight:
Body weight gains were reduced in all surviving animals.
Gross pathology:
Gross pathological examinations at 14 days post administration (terminal necropsies) revealed no test substance-dependent findings. The macroscopic changes observed were attributable to the sacrificing procedure or to minor variations which often occur spontaneously in mice of this strain. However, all dose animals killed in extremis or died spontaneously revealed alterations which were considered to be test substance-related.
Details of necropsy findings were provided in the table 2 under ‘Any other information on results incl. tables’.

Any other information on results incl. tables

Table 1: Cumulative table of pre-terminal deaths (Study# 71231)

Group

Dose (mg/kg)

No. of animals

Post-treatment time

24 hours

7 days

14 days

m

f

m

f

m

f

I

125

5m + 5f

0/5

0/5

1/5

1/5

1/5

1/5

II

250

5m + 5f

0/5

0/5

2/5

1/5

2/5

1/5

III

500

5m + 5f

1/5

0/5

4/5

3/5

4/5

3/5

IV

750

5m + 5f

1/5

0/5

3/5

5/5

3/5

5/5

V

1000

5m + 5f

1/5

1/5

5/5

4/5

5/5

4/5

 

Table 2: Summary of Necropsy findings (Study# 71231)

Dose group

Time point post dose

Specific findings

125 mg/kg

72 hours

Hyperemic and dilated stomach and intestine, pale discoloration of spleen, reduced size of spleen, hyperemic kidney

14 days

Pale discoloration of kidney, thick skinned surface hyperemic pelvis

250 mg/kg

48 hours

None

14 days

Dark discoloration of kidney

500 mg/kg

24-48 hours

Blue discoloration and hyperemic stomach, hyperemic intestine, dark discoloration of liver, pale discoloration and reduced size of spleen, hyperemic kidney, hydrometra of genital system

14 days

Hyperemic kidney

750 mg/kg

24-72 hours

Blue discoloration and hyperemic stomach, dilated intestine, pale discoloration of spleen, pale discoloration and hyperemic pelvis

14 days

Pale discoloration of liver, spleen, kidney

1000 mg/kg

24-48 hours

Blue discoloration and hyperemic stomach and intestine, dark discoloration of liver, pale discoloration of spleen, redden and blue dark discoloration of pelvis

14 days

Pale discoloration of kidney

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
Acute oral administration of Lehmannblau sulfat (2- amino-4-hydroxyethylaminoanisole sulfate) to male and female mice resulted in a LD50 value of 327 mg/kg (Male LD50: 333 mg/kg; female LD50: 351 mg/kg). The registered substance is classified as Category 4 according to GHS regulation.
Executive summary:

The purpose of this study was to determine the acute oral toxicity (LD50) of Lehmannblau sulfat (2- amino-4-hydroxyethylaminoanisole sulfate) when administered once orally to male and female mice.

BOR: NMRI white mice were used in this study. Test substance was administered by gavage using a stomach tube at dose levels of 125, 250, 500, 750 and 1000 mg/kg bw. The dose levels were based on the results of a range finding study where a dose of 2000 mg/kg bw was administered to 2 female animals and both animals died 24 hours post dosing.

Clinical observations were conducted at regular intervals during the 14-day observation period. Body weights were measured at Days 0, 7 and 14. Animals found dead or killed in extremis were immediately necropsied.

The test substance caused dose related clinical signs increasing in severity to CNS, coordination, reflexes and autonomic functions up to 72 hours after dosing. Weight gains were reduced in all surviving animals.

Out of 10 animals (5 males and 5 females), only 1 female survived at 1000 mg/kg bw. At 750 mg/kg bw, only 2 male animals survived. One male animal and 2 female animals survived at 500 mg/kg bw. At 250 mg/kg bw, the mortality rate was low and 4 female and 3 male animals survived. 4 animals/sex survived at 125 mg/kg. Gross pathological examinations at 14 days post administration (terminal necropsies) revealed no test substance-dependent findings in any of the dose groups.

Based on the observed mortality, the following LD50 figures were determined at 14 days according to Finney:

LD50 mouse, female: 351 mg/kg bw

LD50 mouse, male: 333 mg/kg bw

LD50 mouse male + female: 327 mg/kg bw

Acute oral administration of Lehmannblau sulfat (2- amino-4-hydroxyethylaminoanisole sulfate) to male and female mice resulted in a LD50 value of 327 mg/kg  (Male LD50: 333 mg/kg; female LD50: 351 mg/kg).  The registered substance is classified as Category 4 according to GHS regulation.