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Administrative data

Description of key information

acute oral toxicity: LD 50= 750 mg/ kg bw; 
acute inhalative toxicity: LD50= 862 ppm;
acute dermal toxicity: LD50=3250 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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions (no necropsy and no histopathological examinations were performed)
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Deviations:
yes
Remarks:
Analytical purity not reported. Females were not tested. Fasting of animals not reported. No necropsy and no histopathological examinations. Body weights not recoreded. Few informations on environmental conditions.
GLP compliance:
no
Remarks:
GLP was not mandatory at the time of the study
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
other: Wistar-II
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Winkelmann, Germany
- Age: 6 - 7 weeks
- Weight at study initiation: 160 -180 g
-Housing conditions. animals were housed in groups of 5 per cage


Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
single oral application of different doses of undiluted substance to groups of male rats
Doses:
0.600 - 0.650 - 0.700 - 0.800 - 0.900 mL/kg bw, undiluted (corresponding to 600, 650, 700, 800, 900 mg/kg bw (density: 0.998 g/cm³)
No. of animals per sex per dose:
10
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Necropsy of survivors performed: no
- Other examinations performed: clinical signs
Statistics:
Calculation of the median lethal dose (LD50) according to Fink and Hund (Arzneim. Forsch. 15, 1965, 624)
Sex:
male
Dose descriptor:
LD50
Effect level:
750 mg/kg bw
95% CL:
710 - 790
Mortality:
Dose [ml/kg bw]: 600 mg/kg: 0/10 650 mg/kg: 1/10 4d post application 700 mg/kg: 4/10 within 3-4 days post application 800 mg/kg: 6/10 within 2-4 days post application 900 mg/kg: 10/10 within 2-5 days post application
Clinical signs:
600 mg/kg: reduced general condition at day 2; 650 mg/kg: reduced general condition, anesthesia day 1 up to day 14; 700 mg/kg: reduced general condition, anaesthesia, increased diurese - day 1 up to day 14; 800 mg/kg: reduced general condition, anaesthesia, cyanosis, bloody eyes - day 1 up to day 14; 900 mg/kg: reduced general condition, anaesthesia, cyanosis, bloody eyes - day 1 up to time of death
MORTALITY:
Dose [ml/kg bw]:
600 mg/kg: 0/10
650 mg/kg: 1/10 4d post application
700 mg/kg: 4/10 within 3-4 days post application
800 mg/kg: 6/10 within 2-4 days post application
900 mg/kg: 10/10 within 2-5 days post application

CLINICAL SIGNS:
600 mg/kg: reduced general condition at day 2
650 mg/kg: reduced general condition, anesthesia
day 1 up to day 14
700 mg/kg: reduced general condition, anaesthesia,
increased diurese - day 1 up to day 14
800 mg/kg: reduced general condition, anaesthesia,
cyanosis, bloody eyes - day 1 up to day 14
900 mg/kg: reduced general condition, anaesthesia,
cyanosis, bloody eyes - day 1 up to time of
death
Executive summary:

Löser E (1978)

In male Wistar rats (n = 10) dosed with 0.6, 0.65, 0.7, 0.8 or 0.9 ml/kg bw (app. 600, 650, 700, 800 or 900 mg/kg bw) undiluted o-toluidine, deaths occurred at dose levels of = 0.65 ml/kg bw within 2 - 5 days after application. In moribund animals anesthesia, increased diurese, cyanosis, bloody eyes were observed. No necropsy and no histopathological examinations were performed. From this study a LD50 value of 0.75 ml/kg bw (corresponding to 750 mg/kg bw) was derived. This study was conducted with a method similar to OECD guideline 401 with acceptable deviations (Analytical purity not reported. Females were not tested. Fasting of animals not reported. No necropsy and no histopathological examinations. Body weights not recoreded. Few informations on environmental conditions of the animals.)

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
750 mg/kg bw
Quality of whole database:
There is a reliable study which is performed equal to the current guidelines and therefore evaluated with Klimisch score 2

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Although only adopted from OECD SIDS Report published by UNEP basic data are given and significant symptoms are reported to evaluate this toxicological endpoint
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Principles of method if other than guideline:
LC50 was calculated with the Probit Analysis, Finney DJ, 3rd ed. Cambridge University Press.
GLP compliance:
not specified
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Age at study initiation: 8 weeks
- Weight at study initiation: 230-260 g



Route of administration:
inhalation
Type of inhalation exposure:
head only
Vehicle:
other: air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE
The atmosphere was generated by passing nitrogen over o-toluidine liquid contained in a 3-neck round-bottom flask heated from 115 to 120°C. The vapor/aerosol was diluted with humified and O2-enriched houseline air and passed into the exposure chamber. Test atmosphere was regularly controlled.
Duration of exposure:
4 h
Concentrations:
492, 606, 722, 799, 848, 931, 1000 ppm (approx. 2184, 2691, 3206, 3548, 3765, 4134, 4440 mg/m³)
No. of animals per sex per dose:
10
Control animals:
other: not applicable
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: body weights and clinical signs were observed daily
Statistics:
LC50 calculated with the Probit Analysis, Finney DJ, 3rd ed. Cambridge University Press.
Sex:
male
Dose descriptor:
LC50
Effect level:
862 ppm
95% CL:
816 - 913
Exp. duration:
4 h
Remarks on result:
other: ca. 3827 mg/m³: clinical signs included cyanosis, labored breathing, lethargy
Mortality:
492 ppm: 0/10
606 ppm: 0/10
722 ppm: 1/10
799 ppm: 2/10
848 ppm: 6/10
931 ppm: 5/10
1000 ppm: 10/10
Clinical signs:
other: Compound related signs of intoxication, observed during and immediately following exposure, consisted of tremor, slight to moderate cyanosis, muscle spasm, labored breathing, slight to moderate corneal opacity, prostation and semi- prostation, and redd
Body weight:
Body weight losses of 6 to 22 % were observed 1 to 3 days post exposure, with normal weight gains occurring thereafter (individual animal data not given).
Executive summary:

DuPont Chem, 1981, cited in OECS SIDS for o-toluidine in 2005, reported of groups of ten male rats that were exposed head-only to o-toluidine-vapor/aerosol for 4 hours in concentrations ranging from 492 up to 1000 ppm (corresponding to 2184 - 4440 mg/m3). Death occurred at 722 ppm (3206 mg/m3) in 1/10 rats on day 3 post exposure and the concentration of 1000 ppm (4440 mg/m3) was lethal to all animals within 24 hours post exposure. Major treatment related clinical signs included cyanosis, labored breathing, lethargy, prostration, reddish-brown nasal discharge and stained wet perinea, symptoms which show clear evidence for methemoglobinemia. In this study no histopathological investigations were performed. The calculated LC50 value is 862 ppm (3827 mg/m³).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LC50
Value:
3 827 mg/m³
Quality of whole database:
A peer reviewed publication describes this inhalation study providing information that leads to evaluation with Klimisch Score 2

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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given (No data on mortality, no data on clinical signs, no necropsy and no histopathological examinations were performed/reported)
Principles of method if other than guideline:
according to Draize et al., J.Pharmacol.Exper.Therap. 82, 377 (1944)
GLP compliance:
no
Remarks:
GLP was not mandatory at the time of the study
Test type:
standard acute method
Species:
rabbit
Strain:
New Zealand White
Sex:
male
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
-Type of wrap: plastic film
Duration of exposure:
24 h
Doses:
No data on the doses employed were given in the publication. However, the publication stated that doses above 20 mL/kg could not be retained in contact with the skin of the animals

No. of animals per sex per dose:
4
Control animals:
no
Details on study design:
Penetration of rabbit skin was estimated by a technique closely alike to the one-day cuff method of Draize and associates. Before application, the fur was clipped from the entire trunk of the animals and the substance applied under a plastic film for 24h within which time the animals were immobilized (type of wrap: occlusive). The film was removed at the end of the application period and the animals were subsequently caged and observed for another 14d. LD 50 and its fiducial range were estimated by the method of Thompson (Use of Moving averages and interpolation to Estimate Median Effective Dose. Bacteriol. Rev. 1 115 (June 1947)) using the tables of Weils (Tables for Convenient Calculation of Median-Effective Dose (LD50 or: ED 50) and Instructions in Their Use. Biometrics 8: 249 (Sept. 1952))

Statistics:
LD 50 and its fiducial range were estimated by the method of Thompson (Use of Moving averages and interpolation to Estimate Median Effective Dose. Bacteriol. Rev. 1 115 (June 1947)) using the tables of Weils (Tables for Convenient Calculation of Median-Effective Dose (LD50 or: ED 50) and Instructions in Their Use. Biometrics 8: 249 (Sept. 1952))
Sex:
male
Dose descriptor:
LD50
Effect level:
3 250 mg/kg bw
95% CL:
2 010 - 5 250
Mortality:
No data on mortality
Clinical signs:
No data on clinical symptoms
Body weight:
No data on body weights of the animals at test termination
Gross pathology:
No data on necropsy

Dermal application of 2010 - 5250 ml/kg bw over a period of 24 hours followed by a 14-day observation period yielded a LD50 value of 3250 mg/kg bw using groups of 4 male rabbits and the one-day cuff method of Draize. Signs of toxicity and individual animal data were not reported.
Executive summary:

Penetration of rabbit skin was estimated by a technique closely alike to the one-day cuff method of Draize and associates using groups of 4 male rabbits. Before application, the fur was clipped from the entire trunk of the animals and the substance (2010 -5250 ml/kg bw) applied under a plastic film for 24h within which time the animals were immobilized (type of wrap: occlusive). The film was removed at the end of the application period and the animals were subsequently caged and observed for another 14d. LD50 value of 3250 mg/kg bw was calculated

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
3 250 mg/kg bw
Quality of whole database:
This is a reliable study performed by a method equal to the respective guideline and therefore evaluated with Klimisch score 2

Additional information

OECD SIDS

Studies in Animals

Inhalation

Groups of ten male rats were exposed head-only to o-toluidine-vapor/aerosol for 4 hours in concentrations ranging from 492 up to 1000 ppm (corresponding to 2184 - 4440 mg/m3). Death occurred at 722 ppm (3206 mg/m3) in 1/10 rats on day 3 post exposure and the concentration of 1000 ppm (4440 mg/m3) was lethal to all animals within 24 hours post exposure. Major treatment related clinical signs included cyanosis, labored breathing, lethargy, prostration, reddish-brown nasal discharge and stained wet perinea. In this study no histopathological investigations were performed. The calculated LC50value is 862 ppm (3827 mg/m3, DuPont Chem, 1981).

 

Dermal

Dermal application of 2010 - 5250 ml/kg bw over a period of 24 hours followed by a 14-day observation period yielded a LD50value of 3250 mg/kg bw using groups of 4 male rabbits and the one-day cuff method of Draize (Smyth et al., 1962). Signs of toxicity and individual animal data were not reported. In a further study with female Wistar rats single dermal application of 0.75, 1, or

1.25 % o-toluidine-solution (dosing volume not mentioned) resulted in dose-related increase in

methemoglobinemia up to 8 - 10 % (Senczuk and Rucinska 1984).

 

Oral

There are no studies according to the current OECD Test Guidelines, but there is one study, which is adequately documented and is considered of sufficient quality to allow an evaluation of this endpoint. In male Wistar rats (n = 10) dosed with 0.6, 0.65, 0.7, 0.8 or 0.9 ml/kg bw (app. 600, 650, 700, 800 or 900 mg/kg bw) undiluted o-toluidine deaths occurred at dose levels of = 0.65 ml/kg bw within 2 - 5 days after application. In moribund animals anesthesia, increased diurese, cyanosis, bloody eyes were observed. No necropsy and no histopathological examinations were performed.

From this study a LD50value of 0.75 ml/kg bw (corresponding to 750 mg/kg bw) was derived (Bayer AG, 1978).

Oral administration of 50 mg/kg bw o-toluidine to 2 cats per sex resulted in 59.6 to 71.7 % methemoglobin within 4 hours after administration, lateral position, tachypnea, cyanosis mydriasis, apathy, salivation and the death of one cat. During the 14 day-observation period, one cat was in poor condition and 2 cats recovered (BASF AG, 1979).

 

Studies in Humans

Cases of poisoning of workers with o-toluidine following acute exposure are described e.g.

Stark, 1892, cited in Greim (1997), who reported of a worker who inhaled vapor of toluidine (isomer not specified) during the transfer from one open vessel to another one. He lost consciousness, was cyanotic, exhaled levels of toluidine and suffered from strangury and blood was detected in the urine. Full recovery required 5 weeks. In another case, during repair of machines, a fitter was accidentally exposed against o-toluidine (exposure route probably: inhalation, concentration not given). Treatment with tolonium chloride reduced methemoglobinemia from 39.6 % to 2.6 % (BASF AG, 1989). Goldblatt (1955), reported in a survey article that concentrations of 40 ppm (176 mg/m3) toluidine (isomere not specified) in the atmosphere for more than 60 minutes caused severe toxic effects in workers, 10 ppm (44 mg/m3) lead to symptoms of illness and concentration in the atmosphere greater than 5 ppm (22 mg/m3) indicate unsatisfactory conditions (no further details included). In the recent open literature no further cases of acute poisoning are reported.

Conclusion

The LC50(rat) is 852 ppm/4 hrs (approx. 3827 mg/m3/4 hrs), and oral LD50(rat) is 750 mg/kg bw.

The dermal LD50(rabbit) is 3250 mg/kg bw in a limited study. The predominant symptoms after inhalation or oral application were cyanosis, labored breathing, lethargy or loss of consciousness.

o-Toluidine is a methemoglobin forming chemical; this was shown in rats and cats as well as in

humans.


Justification for selection of acute toxicity – oral endpoint
There is a reliable study which is performed equal to the current guidelines and therefore evaluated with Klimisch score 2

Justification for selection of acute toxicity – inhalation endpoint
A peer reviewed publication describes this inhalation study providing information that leads to evaluation with Klimisch Score 2

Justification for selection of acute toxicity – dermal endpoint
This is a reliable study performed by a method equal to the respective guideline and therefore evaluated with Klimisch score 2

Justification for classification or non-classification

The primary toxic effect of o-toluidine is methaemoglobin formation. Taking into account that humans are in general more sensitive to methaemoglobin producing substances than rats o-toluidine is classified as T, Toxic and labelled as R 23/25, toxic by inhalation and if swallowed. Due to the methemoglobin forming property Xn, R21= harmfull in contact with skin should be added. According to Regulation (EC) 1272/2008 the above mentioned statement leads ot tha allocation of o-toluidine to Category 3 (H331 and H301) with respect to acute inhalation and oral exposure and to Category 4 (H312) with respect to dermal exposure.