Registration Dossier

Administrative data

Description of key information

LD50oral > 5000 mg/kg body weight, Wistar rat, 5 males and 5 females, according to EPA OPP 81 -1 (Acute oral toxicity), fixed dose procedure)

LD50dermal > 4000 mg/kg body weight, NZW rabbit, 5 males and 5 females, according to EPA OPP 81 -2 (Acute dermal toxicity), semiocclusive)

LC50 (4h) inhalation: 0.281 mg/L aerosol, Wistar rat, 5 males and 5 females, according to EPA OPP 81 -3 (Acute inhalation toxicity)

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:
1987-11-25 - 1987-12-09
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EPA OPP 81-1 (Acute Oral Toxicity)
Version / remarks:
revised Nov. 1984
Deviations:
no
GLP compliance:
yes
Remarks:
according to US FDA (21 CFR 58) and US EPA (40 CFR 160 and 40 CFR 792)
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
Young adult rats of the Wistar strain (8 weeks old at study start, SPF-quality, randomly bred) were supplied by Charles River Wiga GmbH, Sulzfeld, FRG. Upon receipt each animal was identified with an ear tag. At least five days prior to dosing (acclimatization period) the animals were individually housed in polycarbonate cages containing purified sawdust (Woody clean supplied by Broekman Institute, Someren, The Netherlands) as bedding material. The animals had free access to tap-water (via automatic-nozzles) and a standard pelleted laboratory animal diet (RMH-B, Hope Farms, Woerden, The Netherlands). Certificates of analysis for both diet and drinking water are retained in the N0T0X archives.

ENVIRONMENTAL CONDITIONS
The animal room was air-conditioned, with the temperature maintained within the range of 20-22°C and the relative humidity within the range of 55-70% during the study. The artificial light cycle was 12 hours light, 12 hours dark.
Route of administration:
oral: gavage
Vehicle:
methylcellulose
Details on oral exposure:
Animals were fasted overnight prior to dosing until 3 hours after administration of the test substance. The test substance was suspended in 1% aqueous methyl cellulose (Boom B.V.. Meppel, The Netherlands) and administered once only by gavage using a stainless steel stomach cannula attached to a disposable plastic syringe. The dose level was 5000 mg/kg body weight. Each time the dose volume was 15 ml/kg body weight. The day of dosing was designated as day 0.
Doses:
5000 mg/kg body weight
No. of animals per sex per dose:
5 males
5 females
Control animals:
no
Details on study design:
Observations
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: on the day of dosing (approximately once every two hours) and once daily thereafter for 14 days
- Necropsy of survivors performed: yes
Statistics:
not perfomed
Preliminary study:
In order to establish an appropriate dose range three groups of animals, each comprising 1 male and 1 female, were dosed with an oral dose of test substance at 3200, 4200 and 5600 mg/kg body weight. No mortality occurred during the 7-day observation period. Signs of toxicity were body weight loss, lethargy, piloerection, bloody eye encrustation and diarrhoea. Based on the absence of mortality and the low toxicity, observed in the pilot study, the test could be performed as a limit test. One group of animals, comprising 5 males and 5 females, was treated with a single oral dose of test substance at 5000 mg/kg body weight.
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 5 000 mg/kg bw
Based on:
test mat.
Mortality:
One female rat died on day 4 of dosing.
Clinical signs:
Signs of toxicity were body weight loss for the female rat found dead, lethargy, piloerection, dacryorrhoea, diarrhoea and emaciation.
Body weight:
Signs of toxicity were body weight loss for the female rat found dead. All surviving animals showed body weight gain during the study period (Table 2). However, totally 5 animals revealed body weight loss or stagnant body weight during the first week of observation.
Gross pathology:
Macroscopic examination of animals at termination did not reveal any abnormalities that were considered test substance related, with the exception of a slightly enlarged liver of the female rat which died on day 4.
Other findings:
none

Table 1: Daily Incidence of Mortality and Clinical Observations fo Male and Female Rats Combined Following Acute Oral Dosing of test material
Dose level (mg/kg) Signs of reaction Number showing effects during day of dosing (hours:minutes) or day of observation (day) Total mortality
00:15 02:15 03:35 1 2 3 4 5 6 7 8 9 10 11 12 13 14
5000 No abnomalities 10 0 0 1 3 9 4 3 4 5 8 9 9 9 9 9 9
Lethargy 0 10 10 0 0 1 5 1 0 0 0 0 0 0 0 0 0
Emaciation 0 0 0 0 0 0 0 0 2 2 0 0 0 0 0 0 0
Piloerection 0 6 10 1 7 1 0 6 5 3 1 0 0 0 0 0 0
Diarrhoea 0 0 0 9 0 0 0 0 0 0 0 0 0 0 0 0 0
Dacryorrhoea 0 5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Deads 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 1/10

Table 2: Indivudual body weights and dose volumes acute oral toxicity study of test material in the rat
Dose Level (mg/kg) Animal number Body Weight (g) on: Body weight Gain (g) Day 0-14 Dose Volume (mL)
Day 0* Day 7 Day 14
5000
(Males)
3306 221 220 304 83 3.3
3308 216 224 291 75 3.2
3310 230 221 301 71 3.5
3312 225 251 313 88 3.4
3314 227 227 303 76 3.4
Mean 224 229 302 79
S.D. 5.4 12.8 7.9 6.8
N 5 5 5 5
5000
(Females)
3113 169 174 218 49 2.5
3115 170 178 222 52 2.6
3117 176 - - - 2.6
3119 174 170 219 45 2.6
3121 185 185 252 67 2.8
Mean 175 177 228 53
S.D. 6.4 6.4 16.3 9.6
N 5 4 4 4
* Day of dosing
Interpretation of results:
other: EU GHS criteria not met
Conclusions:
LD50 value for both males and females was noted as exceeding 5000 mg/kg.
Executive summary:

The acute oral toxicity of the test item was evaluated in this single-dose study in rats according to EPA OPP 81 -1 (Acute oral toxicity) and in compliance with GLP ( US FDA 21 CFR 58 and US EPA 40 CFR 160 and 40 CFR 792).

One group of Wistar rats, comprising 5 males and 5 females, was treated with a single oral dose of the test material at 5000 mg/kg body weight. One female rat died on day 4 of dosing. Signs of toxicity were body weight loss for the female found dead, lethargy, piloerection, dacryorrhoea, diarrhoea and emaciation. These signs were considered reversible since as of day 9 no more abnormalities were observed during the 14-day observation period. All animals showed body weight gain during the study period. Macroscopic examination of animals at termination did not reveal any abnormalities that were considered test substance related, with the exception of a slightly enlarged liver of the female which died on day 4. Since only one animal died, the LD50 value for both males and females was noted as exceeding 5000 mg/kg body weight.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
5 000 mg/kg bw
Quality of whole database:
High qualitiy due to well performed test in accordance with GLP and official EPA OPP Guideline.

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
Study period:
1988-02-20 - 1988-03-15
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EPA OPP 81-3 (Acute inhalation toxicity)
Deviations:
no
GLP compliance:
yes
Remarks:
Federal Register, Vol 48, No 230, 1983
Test type:
traditional method
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (U.K.) Limited, Mansion, Kent, England (one batch (16 males and 16 females was received at IRI on 2 February 1988 whilst the second batch (16 males and 16 females) was received on 4 February 1988).Five males and 5 females of the first batch received and 15 males and 15 females of the second batch received were used on the study.
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 5-7 weeks old (arrival)
- Weight at study initiation: approximately 110-135 g (arrival)
- Fasting period before study: None. No water or food during the 4 h exposure period.
- Housing: semi-barrier maintained animal room. The animals were housed in Room C16 of the Rodent Toxicology Complex, at the Elphinstone Research Centre. Rats were housed 5 males or 5 females per cage in suspended polypropylene cages with detachable stainless steel tops and bottoms. All cages were suspended over trays containing absorbent paper
- Diet (e.g. ad libitum): ad libitum. Rat and Mouse (modified) No. 1 Diet SQC Expanded, supplied by Soecial Diet Services limited
- Water (e.g. ad libitum): ad libitum. Each cage was supplied with a polypropylene water bottle (capacity 500 ml) with a rubber washer and melamine cap
- Hygiene: Cages were changed weekly and cage tray papers were changed as required (at approximately 3 day intervals), water bottles were cleaned weekly. Each afternoon, when other work in the room was finished, floors were swept then washed with a disinfectant solution. Once each week walls, benches and racking within the animal room were washed with a disinfectant solution.
Disinfectant used was either 1% Tego from T.H. Goldschmidt and Company Limited, or 3% Hycogen from Hy-co Products (Scotland) Limited.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20°C + 2°C (Temperature extremes recorded were 16°C-24°C)
- Humidity (%): ca 55% (humidity extremes were 30%-54%)
- Photoperiod (hrs dark / hrs light): A 12 h light-dark cycle was controlled by a time switch, light hours being 0700-1900 h.
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
other: snout only
Vehicle:
clean air
Mass median aerodynamic diameter (MMAD):
>= 2.1 - <= 2.9 µm
Remark on MMAD/GSD:
The test material was tested as a particulate aerosol in its original form. The particle size distribution of the dispersed material inside the exposure chamber was estimated twice during the exposure period using a Marple (model 296) Caccade Impactor. The device was positioned on the chamber using a vacuum pump, at a constant rate of 2 litres/min for a recorded time period. The impaction substrates (for each stage) were weighed before and after sampling and the weight increase on each substrate determined to be the mass of particles in the size range of that impactor stage. From the results obtained, the total weight of particles on all stages and filter was summed and the percent particle mass in each size range calculated. The data may be presented graphically as a cumulative size distribution, showing the percent of particle mass smaller than the aerodynamic particle diameter, and the respirable mass fraction determined from the particle size distribution.
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
The test material was tested as a particulate aerosol in its original form. The test atmospheres were generated by means of a Wright dust feeder (Wright, B.M., 1950, J. Sci. Instr., 27, 12). In this electrically operated device, a scraper blade removed the powder at a steady rate from a prepacked canister of test material. The loose powder was removed from the canister by a stream of clean, dry compressed air.
After passing through a jet the air containing the finely dispersed dust passed directly into the exposure chamber.
The aluminium exposure chamber was cylindrical in cross section and had a volume of approximately 41.5 litres. An extract duct from the chamber was connected by way of a high efficiency filter to a metered vacuum system. The exposure chamber was mounted inside an extract cabinet for the protection of operators and the environment.
Compressed air was supplied by means of 2 Broomwade compressors (Type CAR 31) fitted with automatic pressure control switches. These supplied filtered, conditioned, oil-free compressed air for subsequent dilution of the test atmosphere.
The canister of the Wright dust feeder was packed in stages to ensure an even density. The canister was positioned on the Wright dust feeder which was located at the top of the exposure chamber. The exit nozzle of the dust feeder was passed through a rubber diaphragm into the chamber space and the exposure started by switching on the dust feeder and engaging the scraper blade with the powder.
The exposure system was truly dynamic, incorporating a single pass of the freshly generated material. The particles dispersed throughout the chamber and exited through the base to a filtered vacuum line. Chamber air flow rates were monitored continuously and the values recorded at 30 min intervals.
The inhalation exposures were conducted in a room adjacent to the animal holding room. Each animal was removed from its cage and examined for general health status. The ear number was checked, the animal weighed and then loaded into a tapered, polycarbonate restraint tube which fitted onto the exposure chamber and sealed by means of a push fit through a rubber 'o' ring. All animals were exposed on a single tier eliminating exposure variations. Only the animals' snouts were exposed to the test atmosphere. The animals were not allowed access to food or water during the 4 h exposure period.
During exposure the animals were observed at regular intervals for signs of any adverse reactions to treatment. On completion of the 4 h exposure period, the animals were removed from the chamber, unloaded from the restraint tube, returned to their cages in the animal holding room and observed for clinical signs.
The study protocol stated that only 3 groups would be exposed to a range of concentrations of the test material. However, due to the 100% mortality observed following the first 2 concentrations tested, an additional group was included in the study to give a total of 4 groups each comprising 5 male and 5 female rats. The animals were exposed to a range of concentrations of the test material via the inhalation route by snout only exposure for a single continuous 4 h period. The exposures were conducted over the period 20 February -1 March 1988.
The first concentration tested was 1.01 mg/litre, the remaining exposure concentrations were selected from the mortality pattern recorded following this initial concentration.
During the exposure periods the temperature within the exposure chamber was measured by a mercury thermometer located at the animals' breathing zone whilst the relative humidity was monitored using wet/dry bulb mercury thermometers. The chamber temperature and relative humidity were measured at 30 min intervals throughout the exposure periods and ranged from 17°C-22°C and 13%-62% respectively for temperature and humidity. The humidity readings at the higher chamber concentrations tended to be extremely low due to the accumulation of test material on the wet bulb thermometer.

TEST ATMOSPHERE
Chamber concentrations were measured at regular intervals during the exposure periods. The gravimetric method used employed pressed glass fibre filters (Whatman GF7B) placed in a filter holder. The conical input side of the holder was positioned and temporarily sealed in a port in the exposure chamber at the animals' breathing zone. Chamber air was drawn through the filter at a measured rate of 1.0 litre/min using a vacuum pump. The air flow during each sample was controlled by acritical orifice and timed for a suitable period. Each filter was weighed before and after sampling in order to calculate by difference the weight of collected material. The chamber concentration was estimated by further calculation using the sample air volume.
The nominal chamber concentration was estimated using the following equation:

Nominal concentration (mg/litre) = Weight of material used (mg) / Total air flow through chamber /(litres)
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
1.01 (not estimated), 0.52 (3.72), 0.15 (0.83) and 0.31(2.10) mg/L measured (nominal)
No. of animals per sex per dose:
5 males and 5 females per dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: All the rats were weighed immediately before dosing and on Days 2, 3, 4, 7, 10 and 14 post exposure.
- Necropsy of survivors performed: yes (Any gross lesions observed were recorded in descriptive terms, including location(s), size (mm), colour and number. The respiratory tract was subjected to detailed macroscopic examination for signs of irritancy or local toxicity. All organs were examined in situ)
- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other: All the rats were observed for clinical signs at approximately hourly intervals throughout the exposure period, for the first 1-2 h post dosing and thereafter at least once daily during the subsequent 14 day observation period. The onset, intensity and duration of any signs observed were recorded. Lungs: Body Weight Ratio:
The lungs of each animal were removed and weighed to allow calculation of lung: body weight ratio.
Statistics:
not reported
Preliminary study:
not performed
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
0.281 mg/L air
Based on:
test mat.
Remarks:
aerosol
95% CL:
>= 0.207 - <= 0.361
Exp. duration:
4 h
Mortality:
The mortality pattern observed as a result of exposure to the test material was as follows:
Group       Concentration (mg/L)       Mortality
Males        Females     Total %
3              0.15                                  0/5            0/5                0
4              0.31                                  3/5            3/5                60
2              0.52                                  5/5            5/5                100
1              1.01                                  5/5            5/5                100
All mortalities occurred overnight following exposure, or on Day 1 post exposure.
Clinical signs:
other: Marked respiratory depression was observed during exposure for the animals in Groups 1 and 2, exposed to 1.01 or 0.52 mg/L respectively. On return to their cages (following completion of exposure) all animals showed heavy deposition of the test material
Body weight:
All animals which survived exposure to the test material showed a marked body weight loss following exposure. During the subsequent 14 day observation period the animals showed a gradual body weight recovery. The overall body weight gains over the 14 day post exposure observation period were lower then normal.
Gross pathology:
Gross post mortem examinations of the animals that died following exposure to the test material revealed extensive haemorrhage in the lungs and resultant pulmonary oedema.
Other findings:
Lung:Body Weight Ratio:
Lung:body weight ratios for those animals that died as a result of exposure to the test material were markedly increased. In addition, the ratios observed for the surviving animals in Group 3 (exposed to 0.31 mg/L) were marginally increased.

Chamber Atmospheric Conditions

Particle Size Distribution

Estimation of the particle size distribution revealed that the percentage of particles <3.5 µm was 67.0%, 74.2%, 81.9% and 85.1% by weight respectively for Groups 1, 2, 3 and 4.

The mass mean diameter of the aerosol particles generated for Groups 1, 2, 3 and 4 was determined to be 2.9 µm, 2.7 µm, 2.2 µm and 2.1 µm respectively.

Interpretation of results:
other: Category 2 based on EU GHS criteria
Conclusions:
The LC50 value of the test material was calculated to be 0.281 mg/L. The results obtained indicated that an exposure concentration as low as 0.15 mg/L produced toxic effects.
Executive summary:

This study was undertaken to investigate the acute inhalation toxicity of the test material and to determine the median lethal concentration (LC50) in rats following a single 4 hour snout only exposure. The study was conducted in compliance with EPA Guideline Section 81-3 and was in compliance with Good Laboratory Practice Standards (FED. Reg. Vol 48, No. 230, 1983).

Four groups of Sprague Dawley rats consisting each of 5 males and 5 females were exposed to 0.15, 0.31 0.52 and 1.01 mg/L concentration of the test material. No mortality was observed at 0.15 mg/L whereas at concentrations of 0.52 and 1.01 mg/L all animals died. In the exposure group at 0.31 mg/L 3 males and 3 females died.

Marked respiratory depression was observed during exposure at all concentrations. All or some of the following signs were recorded for all groups: hunched/subdued appearance, piloerection, laboured respiration, hypokinesia and ataxia following exposure. The signs showed a dose related response; being more severe in the animals exposed to 0.52 or 1.01 mg/L. Surviving animals showed a body weight loss following exposure. A gradual body weight recovery was recorded over the 14 day post exposure observation period. Gross pathological examination of the animals that died following exposure to the test material revealed extensive haemorrhage in the lungs and resultant pulmonary oedema. Lung:body weight ratios were markedly increased for the premature decents.

The LC50 of the test material was calculated to be 0.281 mg/L with 95% confidence limits of 0.207-0.361 mg/L.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LC50
Value:
281 mg/m³
Quality of whole database:
High qualitiy due to well performed test in accordance with GLP and official EPA OPP Guidelines. One supporting study was conducted also in compliance with GLP and international accepted methods.

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:
1987-11-19 - 1987-12-03
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EPA OPP 81-2 (Acute Dermal Toxicity)
Version / remarks:
revised Nov. 1984
Deviations:
no
GLP compliance:
yes
Remarks:
US FDA (21 CFR 58) and US EPA (40 CFR 160 and 40 CFR 792)
Test type:
fixed dose procedure
Limit test:
yes
Species:
rabbit
Strain:
New Zealand White
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
Adult rabbits of the NZW strain (9-10 weeks old at study start, SPF-quality) were supplied by the Broekman Institute, Someren, The Netherlands. Upon receipt each animal was identified with an ear tag. At least five days prior to dosing (acclimatization period) the animals were individually housed in metal cages (RUCO, Valkenswaard, The Netherlands). The animals had free access to tap-water (via automatic nozzles) and were held on a restricted diet of a standard laboratory animal feed (LK-01) obtained from Hope Farms, Woerden, The Netherlands. Certificates of analysis for both diet and drinking water are retained in the NOTOX archives.

ENVIRONMENTAL CONDITIONS
The animal room was air-conditioned, with the temperature maintained within the range of 20-21°C and the relative humidity within the range of 55-65% during the study. The artificial light cycle was 12 hours light, 12 hours dark.
Type of coverage:
semiocclusive
Vehicle:
methylcellulose
Details on dermal exposure:
Dosing
On the day of dosing the test substance was suspended in 1% aqueous methyl cellulose (Boom B.V., Meppel , The Netherlands) and was evenly spread on a surgical gauze (6x10 cm) that was fixed to successively micropore and flexible bandage (Coban, 3M, St. Paul, Minn., USA). This patch was applied to the clipped area of each animal and fixed with adhesive tape (Sleek, Smith and Nephew, Hull, U.K.). The dose volume was 7.5 ml/kg body weight. The dose level was 4000 mg/kg body weight. The day of dosing was designated as day 0.

Observations
Cage-side observations were performed on the day of dosing (approximately once every two hours) and once daily thereafter for 14 days. Any signs of toxicity were recorded along with the time of onset and duration. Individual body weights were measured weekly. At the end of the study (day 14) all animals were killed with euthesate and subjected to necropsy. In addition, observations regarding changes of the treated skin surface were performed following bandage removal and on day 4, 7, and 14.
Duration of exposure:
24 hours
Doses:
4000 mg/kg bw
No. of animals per sex per dose:
5 males and 5 females
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: daily (Observations regarding changes of the treated skin surface were performed following bandage removal and on day 4, 7, and 14.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other:
Statistics:
not reported
Preliminary study:
Upon request of the sponsor the test could be performed as a limit test. One group of animals, comprising 5 males and 5 females, was treated with a single dermal dose of test substance at 4000 mg/kg body weight.
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 4 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortalities occurred during the 14-day observation period.
Clinical signs:
Six animals showed a closed 3rd eyelid which was red or swollen in some animals (Table 1). The treated skin surface of the animals revealed slight erythema and oedema, which disappeared by the end of the first week (Table 2). In addition the treated surface was black discoloured due to staining by the test substance.
Body weight:
All animals showed body weight gain during the study period (Table 3).
Gross pathology:
Macroscopic abnormalities at necropsy at the end of the observation period included local dark red areas in the lungs, enlarged spleen, accessory spleen tissue in the pancreas, dilated ileum and some petechiae in the dorso-lateral muscles (Table 1).
Other findings:
not reported
Table 1: Individual Animal Observations
Acute Dermal Toxicity study in the rabbit - Test item at 4000 mg/kg
Physical/Clinical Parameter Animal Number/Sex
3202
M
3204
M
3206
M
3208
M
3210
M
3250
F
3252
F
3254
F
3258
F
3260
F
No effects of treatment DPD 14 14 14
3rd eyelid
- closed DPD 2-4 2
- closed and swollen DPD 1
- closed and red DPD 1 1
Dead KIL X X X X X X X X X X
Necropsy findings:
- no gross lesions X X X X X X X
- Lungs local dark red areas X
- Enlarged spleen X
- Accessory spleen tissue in the pancreas X
- diated ileum X
- some small petechiae in dorso-lateral muscles X X
Glossary of terms for lsit of individuel findings:
DPD = Days post dosing
KIL = Killed at necropsy
KGL = no gross lesions
X = positive

Table 2: Group Incidences for irritation of the treated skin surface
Acute dermal toxicity study in the rabbit - Test iteml at 4000 mg/kg body weight
Observation Day of Evaluation
1* 4 7 14
No abnomalities 0 3 5 10
Sligth erythema 9 0 0 0
Oedema 9 2 0 0
Black discolouration 9 6 5 0
* Following bandage removal

Table 3: Indivudual body weights and dose volumes
Acute dermal toxicity study in the rabbit - Test item at 4000 mg/kg body weight
Dose Level (mg/kg) Animal number Body Weight (g) on: Body weight Gain (g) Day 0-14 Dose Volume (mL)
Day 0* Day 7 Day 14
4000
(Males)
3202 2196 2343 2536 340 16.5
3204 2395 2402 2513 118 18.0
3206 1804 1920 2174 370 13.5
3208 2065 2237 2358 293 15.5
3210 1991 2125 2290 299 14.9
Mean 2090 2205 2347 284
S.D. 221.6 191.4 152.4 98.0
N 5 5 5 5
4000
(Females)
3250 2145 2279 2347 202 16.1
3252 2152 2283 2402 250 16.1
3254 2277 2352 2470 193 17.1
3258 2172 2215 2356 184 16.3
3260 2349 2424 2502 153 17.6
Mean 2219 2313 2415 196
S.D. 90.1 83.4 68.7 35.2
N 5 5 5 5
* Day of dosing
Interpretation of results:
other: EU GHS criteria not met
Conclusions:
Under the conditions used in this study, the LD50 value for both males and females was noted as exceeding 4000 mg/kg.
Executive summary:

The test material has been assessed in an acute dermal toxicity study in accordance with U.S. EPA FIFRA Guideline 81 -2 and in compliance with GLP. One group of NZW rabbits, comprising 5 males and 5 females, was treated with a single dermal dose of the test item at 4000 mg/kg body weight for 24 hours. No mortalities occurred during the 14-day observation period. A sign of toxicity was a closed and red or swollen 3rd eyelid. The treated skin surface of the animals revealed slight erythema and oedema, which disappeared by the end of the first week. In addition the treated surface was black discoloured due to staining by the test substance. All animals showed body weight gain during the study period. Macroscopic abnormalities noted in animals at necropsy included local dark red areas in the lungs, enlarged spleen, accessory spleen tissue in the pancreas, dilated ileum and petechiae in the dorso-lateral muscles. The LD50 value for both males and females was noted as exceeding 4000 mg/kg body weight.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
4 000 mg/kg bw
Quality of whole database:
High qualitiy due to well performed test in accordance with GLP and official EPA OPP Guideline.

Additional information

Acute toxicity: oral

The acute oral toxicity of the test item was evaluated in this single-dose key study (Rejinders, 1987a) in rats according to EPA OPP 81 -1 (Acute oral toxicity) and in compliance with GLP ( US FDA 21 CFR 58 and US EPA 40 CFR 160 and 40 CFR 792).

One group of Wistar rats, comprising 5 males and 5 females, was treated with a single oral dose of the test material at 5000 mg/kg body weight. One female rat died on day 4 of dosing. Signs of toxicity were body weight loss for the female found dead, lethargy, piloerection, dacryorrhoea, diarrhoea and emaciation. These signs were considered reversible since as of day 9 no more abnormalities were observed during the 14-day observation period. All animals showed body weight gain during the study period. Macroscopic examination of animals at termination did not reveal any abnormalities that were considered test substance related, with the exception of a slightly enlarged liver of the female which died on day 4. Since only one animal died, the LD50 value for both males and females was noted as exceeding 5000 mg/kg body weight.

Acute toxicity: dermal

The test material has been assessed in an acute dermal toxicity study (Rejinders, 1987b) in accordance with U.S. EPA FIFRA Guideline 81 -2 and in compliance with GLP. One group of NZW rabbits, comprising 5 males and 5 females, was treated with a single dermal dose of the test item at 4000 mg/kg body weight for 24 hours. No mortalities occurred during the 14-day observation period. A sign of toxicity was a closed and red or swollen 3rd eyelid. The treated skin surface of the animals revealed slight erythema and oedema, which disappeared by the end of the first week. In addition the treated surface was black discoloured due to staining by the test substance. All animals showed body weight gain during the study period. Macroscopic abnormalities noted in animals at necropsy included local dark red areas in the lungs, enlarged spleen, accessory spleen tissue in the pancreas, dilated ileum and petechiae in the dorso-lateral muscles. The LD50 value for both males and females was noted as exceeding 4000 mg/kg body weight.

Acute toxicity: inhalative

The key study (McDonald, 1988) was undertaken to investigate the acute inhalation toxicity of the test material and to determine the median lethal concentration (LC50) in rats following a single 4 hour snout only exposure. The study was conducted in compliance with EPA Guideline Section 81-3 and was in compliance with Good Laboratory Practice Standards (FED. Reg. Vol 48, No. 230, 1983).

Four groups of Sprague Dawley rats consisting each of 5 males and 5 females were exposed to 0.15, 0.31 0.52 and 1.01 mg/L concentration of the test material. No mortality was observed at 0.15 mg/L whereas at concentrations of 0.52 and 1.01 mg/L all animals died. In the exposure group at 0.31 mg/L 3 males and 3 females died.

Marked respiratory depression was observed during exposure at all concentrations. All or some of the following signs were recorded for all groups: hunched/subdued appearance, piloerection, laboured respiration, hypokinesia and ataxia following exposure. The signs showed a dose related response; being more severe in the animals exposed to 0.52 or 1.01 mg/L. Surviving animals showed a body weight loss following exposure. A gradual body weight recovery was recorded over the 14 day post exposure observation period. Gross pathological examination of the animals that died following exposure to the test material revealed extensive haemorrhage in the lungs and resultant pulmonary oedema. Lung:body weight ratios were markedly increased for the premature decents.

The LC50 of the test material was calculated to be 0.281 mg/L with 95% confidence limits of 0.207-0.361 mg/L.

In a supporting study (Hardy, 1988) the acute inhalation toxicity of the test material was assessed by exposing 4 groups of Albino rats (5 males and 5 females per group) each (whole body) for a continous period of 4 hours to aerosols produced from the test substance and followed the test guidelines according to EPA FIFRA 81-3, EPA TOSCA 798-1150, OECD Guideline 403, EU Method B2 and in compliance wit GLP (US EPA CFR 40 Part 160, 1983). The observation period was 14 days and revealed the following mortalities:

Level (mg/L) Mortalities
Males Females Total
0.336 0/5 1/5 1/10
0.414 1/5 5/5 6/10
0.548 5/5 5/5 10/10
0.21 5/5 5/5 10/10

During exposure clinical signs were consistend with exposure to an irritant dust including partial closing of the eyes, fluid discharge from the mouth or nose, exaggerated respiratory movements and the adoption of a hunched posture. During the obervation period the signs seen immediately following exposure to the test materal included disturbances to respiration and lethargy. Abnormal respiration persisted for up to 12 days following exposure. All rats were normal at the end of the observation period. There were decreases of bodyweight or reductions in the rate of bodyweight gain for up to 4 days following exposure. Subsequently weight gain for rats that survived exposure to the test material was similar to that of the control rats.

The findings for rats that died as a result of exposure to the test material were typified by congestion of the lungs and the presence of fluid in the trachea. There were no treatment-related macroscopic abnormalities in rats that survived exposure to the test material. The lung weight to bodyweight ratio was markedly increased, due to a high lung weight in rats that died as a result of exposure to the test material. The lung weights for some (ca. 50%) rats that survived exposure to the test material were marginally higher than control values. The lung weights were within normal limits for the control rats and for some rats that survived exposure to the test material

The LC50 (4-hour) for the test item is estimated at 0.40 mg/L of air. The standard error of the estimate is 0.016 mg/L.

Justification for classification or non-classification

The key studies for oral acute toxicity of the test item reveal an acute oral LD50 of greater than 5000 mg/kg bw in rats. The key study for dermal acute toxicity of the test item shows an acute dermal LD50 of greater than 4000 mg/kg bw in rats.

The key study for inhalation acute toxicity of the test item shows an acute inhalation LC50(4h) of 0.281 mg/L (aerosol) in rats. This finding is supported by Hardy, 1988, showing an LC50(4h) of 0.40 mg/L (aerosol) in rats.

Therefore the test item does not meet the criteria for classification and labelling for oral and dermal toxicity in accordance with the Regulation (EC) No 1272/2008 (CLP Regulation). The test item does meet the the criteria for classification and labelling for inhalation toxicity as Category 2 in accordance with the Regulation (EC) No 1272/2008 (CLP Regulation).