2-Pyridinecarboxylic acid, 4-amino-3,6-dichloro-

Endpoint summary

Description of key information

ORAL
LD50 > 5000 mg/kg bw/day, Fischer 344 rat (male/female), OECD 401, EU Method B.1, EPA OPPTS 870.1100, JMAFF (2000), Brooks & Yano (2001)
LD50 > 5000 mg/kg bw/day, Wistar rat (female), OECD 423, EU Method B.1, EPA OPPTS 870.1100, JMAFF 2 -1 -1, Patel (2018)

INHALATION
LC50 > 5.5 mg/L, Fischer 344 rat (male/female), OECD 403, EU Method B.2, EPA OPPTS 870.1300, JMAFF Nousan 4200, Kiplinger (2001)
LC50 > 5.49 mg/L, Wistar rat (male/female), OECD 403, EU Method B.2, EPA OPPTS 870.1300, JMAFF 2 -1 -3, Patel (2018)

DERMAL
LD50 > 5000 mg/kg bw/day, Fischer 344 rat (male/female), OECD 402, EU Method B.3, EPA OPPTS 870.1200, JMAFF (2000), Brooks & Yano (2001)

LD50 > 5000 mg/kg bw/day, Wistar rat (female), OECD 402, Patel (2018)

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 June 2001 to 30 August 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 (Acute Toxicity (Oral))

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF Acute Oral Toxicity Study, 2000

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

Purity: 94.5%

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: Approximately 11 weeks
- Weight at study initiation: 130 - 291 g
- Fasting period before study: rats were fasted the night prior to treatment. Feed was provided to all rats immediately following administration of the test material.
- Housing: Animals were housed two or three per cage in stainless steel cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: At least one week

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 3 °C
- Humidity: 40 - 70 % (relative)
- Air changes: 12 - 15 air changes per hour
- Photoperiod: 12 hours of darkness / 12 hours of light (06:00 to 18:00)

Route of administration:

oral: gavage

Vehicle:

methylcellulose

Details on oral exposure:

Five male and five female Fischer 344 rats received the test material at a dose of 5000 mg/kg body weight as a 50 % mixture in 0.5 % aqueous methylcellulose in two fractional doses approximately one hour apart.

Doses:

5000 mg/kg as a 50 % mixture in vehicle

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: A Detailed Clinical Observation (DCO) was conducted for all rats prior to test material administration for comparison with the observations recorded throughout the study. Animals were observed a minimum of two times on the day of treatment. A DCO was done each day (including weekends and holidays) during the study. Hand-held and open-field observations included a careful physical examination. Each animal was weighed pre-study, on the day of treatment, and on test days 2, 8, and 15.
- Necropsy of survivors performed: Yes, necropsy was performed on all animals. Animals were anaesthetised by inhalation of carbon dioxide and euthanised by decapitation after clamping of the trachea.
- Other examinations performed: The eyes were examined in situ using a moistened glass microscope slide applied to the corneal surface. Following inspection of the externum and body orifices, the nasal, cranial, oral, thoracic, and abdominal cavities were opened and the visceral organs were examined both in situ and following dissection, and tissues were not saved.

Statistics:

Means and standard deviations were calculated for body weights. The data were evaluated for statistical outliers by a sequential test (Grubbs, 1969); however, outliers were not routinely excluded from statistical analysis.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Mortality:

One male died on study day 3. All other animals survived the duration of the study.

Clinical signs:

other: Clinical observations for the male that died were consistent with the rat’s moribund condition. Clinical observations of high incidence for the surviving rats consisted of various combinations of: perineal soiling (9 of 9 rats), watery faeces (7 of 9 rats

Gross pathology:

The male rat that died had treatment-related gross findings consisting of haemolysed blood and gas in the gastrointestinal tract and perineal soiling. The haemolysed blood was consistent with a stress-induced alteration. Surviving animals had no treatment-related gross pathologic observations.

Table 1: Mean body weights

Sex	Days on test
	-1	1	2	8	15
Male	276.2	257.8	248.1	269.6	293.4
Female	151.9	139.6	141.0	152.6	163.0

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this study, the acute oral LD50 of the test material in male and female Fischer 344 rats was greater than 5000 mg/kg.

Executive summary:

The acute oral toxicity of the test material was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 401, EU Method B.1, EPA OPPTS 870.1100 and JMAFF (2000).

During the study, five male and five female Fischer 344 rats received test material at a dose of 5000 mg/kg body weight as a 50 % mixture in 0.5 % aqueous methylcellulose by single dose gavage. Parameters evaluated during the two-week observation period included body weights, detailed clinical observations, and gross pathological changes.

Under the conditions of this study, the acute oral LD50 of the test material in male and female Fischer 344 rats was greater than 5000 mg/kg.

Reference 2

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 (Acute Toxicity (Oral))

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF 2-1-1

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Specific details on test material used for the study:

TSN314667
Batch: ENBK-169021-016
Purity: 95.9%

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Animal Breeding Facility, Jai Research Foundation
- Females (if applicable) nulliparous and non-pregnant: [yes/no] yes
- Age at study initiation: 8 to 9 weeks
- Weight at study initiation: (g) Minimum: 176.2, Maximum: 185.4
- Fasting period before study: overnight fasting prior to dosing and three hours post-dosing
- Housing: Polypropylene rat cages covered with stainless steel grid top were used. Autoclaved clean rice husk was used as the bedding material. Wooden block was provided as enrichment material.
- Diet (e.g. ad libitum): ad libitum except during fasting
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 to 9 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 23 °C
- Humidity (%): 49 to 66%
- Air changes (per hr): Minimum 15 air changes/hour
- Photoperiod (hrs dark / hrs light): 12 h artificial light and 12 h darkness, light hours being 06:00 - 18:00 h.

Route of administration:

oral: gavage

Vehicle:

methylcellulose

Details on oral exposure:

The test item was administered, following overnight fasting, at a dose of 5000 mg aminopyralid TGAI/kg body weight, using 0.5% carboxymethyl cellulose as a vehicle and at a constant dose volume (10 mL/kg).

Doses:

5000 mg/kg

No. of animals per sex per dose:

3

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: 0.5, 1, 2, 3, 4 and 6 hours post-administration on the day of dosing. Subsequently, the rats were observed twice a day for morbidity and mortality for a period of 14 days following oral dosing. The clinical signs were recorded once a day. Individual body weight was recorded prior to dosing on day 0 and on days 7 and 14.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other:mortality, clinical signs and body weights, gross necropsy

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality was observed in female rats treated with 5000 mg aminopyralid TGAI/kg body weight.

Clinical signs:

other: No signs of toxicity were observed in rats treated at the dose level of 5000 mg aminopyralid TGAI/kg body weight.

Gross pathology:

External examination of terminally sacrificed female rats did not reveal any abnormality of pathological significance. Visceral examination of female rats sacrificed at termination did not reveal any lesion of pathological significance.

Interpretation of results:

GHS criteria not met

Conclusions:

Acute oral LD50 (female Wistar rats) > 5000 mg/kg

Executive summary:

The acute oral toxicity of aminopyralid TGAI was investigated using the acute toxic class method. Female Wistar rats (RCCHan:WIST), (approximately 8-9 week old and weighting 176.2-185.4 g at dosing)were given a single oral dose of aminopyralid TGAI. The test item was administered, following overnight fasting, at a dose of 5000 mgaminopyralid TGAI/kg body weight, using 0.5% carboxymethyl cellulose as a vehicle and at a constant dose volume (10 mL/kg).Initially, one female rat was given a single dose of 5000 mg aminopyralid TGAI/kg body weight. As no mortality was observed up to approximately 48 hours at this dose level, another two female rats were administered the same dose level. As no mortality was observed at this dose level further testing was not required. Following dosing, animals were observed for 14 days (in which mortality, clinical signs and body weights were recorded) after which were sacrificed and subjected to gross necropsy examination.

 

No mortality or treatment related clinical signs were observed in any animal throughout the study period.

 

Changes in body weight were considered within the expected range for this strain and age of animals and not influenced by the treatment.

 

No internal or external abnormality was recorded at the post-mortem examination.

 

In conclusion, based on the study results, the acute oral median lethal dose (LD₅₀) of aminopyralid TGAI in female Wistar rats was found to be greater than 5000 mg/kg body weight.

 

Based on the results of this study, an indication of the classification foraminopyralid TGAI is as follows:

 

Globally Harmonized System of Classification and Labelling of Chemicals (GHS 2017): Unclassified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

5 000 mg/kg bw

Quality of whole database:

Two studies were conducted under GLP conditions and in accordance with standardised guidelines; the studies were therefore assigned a reliability score of 1 in line with the criteria of Klimisch et al. (1997). The overall quality of the database is good.

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 May 2001 to 13 June 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1300 (Acute inhalation toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.2 (Acute Toxicity (Inhalation))

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF, Nousan No. 4200

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

Purity: 94.5%

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: Approximately 7 and 9 weeks for males and females, respectively
- Weight at study initiation: 142 - 162 g (on the day of exposure)
- Housing: individually in suspended wire mesh cages
- Diet: ad libitum
- Water: reverse osmosis-treated municipal water, ad libitum
- Acclimation period: The animals were acclimated to laboratory conditions for a minimum of seven days prior to initiation of exposure. The animals for use on study were acclimated to nose-only restraint tubes for a single two-hour period prior to exposure.

ENVIRONMENTAL CONDITIONS
- Temperature: 70 - 73 F
- Humidity: 42 - 49 %
- Photoperiod: 12 hours of darkness / 12 hours of light

Route of administration:

other: inhalation: dust aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

CHAMBER DESCRIPTION
The exposure was conducted in an 8.6-L conventional nose-only exposure system (designed and fabricated by the testing facility). Animals were restrained in nose-only tubes during exposure. Food and water were withheld during the exposure.

TEST ATMOSPHERE GENERATION
The test material was metered at 1.1 g/min by an auger-type dust feeder (AccuRate, Inc., Whitewater, WI) to a jet mill air micronizer (Model 00 Jet-O-Mizer, Fluid Energy Aljet, Plumsteadville, PA) operating as a dispersion device. The dust feeder was equipped with a magnetic vibrator (Syntron Model V-2-8, FMC Corp., Homer City, PA) to facilitate delivery of the test material. The aerosol from the jet mill was piped to the exposure system inlet via ¾-inch Tygon® tubing. Humidified dilution air was added to maintain the protocol-specified humidity range and to further dilute the atmosphere as necessary. Exhaust aerosol was filtered (particulate) prior to entering the in-house exhaust system.

CHARACTERISATION OF EXPOSURE ATMOSPHERES
- Mean temperature: 21 °C
- Mean relative humidity: 30 %

TEST ATMOSPHERE
- Nominal concentrations: The nominal exposure concentration was calculated as the total amount of test material used during the exposure divided by the total volume of air that passed through the system during the exposure.
- Actual concentrations: Actual exposure concentrations were determined by standard gravimetric methods. Samples were collected on pre-weighed 25-mm glass-fibre filters (Type A/E, Gelman Sciences, Ann Arbor, MI) held in an open-face filter holder positioned in the sampling port of the nose-only system. Following sample collection, the filters were re-weighed and the concentration calculated as the filter weight difference (post-sample minus pre-sample) divided by the sample volume.

TEST ATMOSPHERE (if not tabulated)
- Particle size determination: Two aerosol particle size determinations were conducted using a seven-stage cascade impactor (In-Tox Products, Albuquerque, NM). Pre-weighed 25-mm glass-fibre filters (Gelman Sciences) were used as the collection substrates. The samples were collected at 1.8 LPM for 1.5 minutes. One sample was collected during the first two hours and one sample was collected during the last two hours. The filters were reweighed and the particle size calculated based on the impactor stage cut-offs. The aerosol size was expressed as the mass median aerodynamic diameter (MMAD) and the geometric standard deviation (GSD).
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): The exposure aerosol was characterised by a mass median aerodynamic diameter of 2.5 µm with a geometric standard deviation of 2.45.

Analytical verification of test atmosphere concentrations:

yes

Remarks:

by standard gravimetric methods

Duration of exposure:

4 h

Concentrations:

5.5 mg/L

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:
> Mortality: Each rat was observed for mortality approximately hourly during exposure, immediately following exposure on day 0 and twice daily thereafter for 14 days.
> Clinical Observations: Each rat was observed immediately following exposure on day 0 and once daily thereafter for 14 days. Due to the limited visibility of the animals in the restraint tubes, observations during exposure were limited to evaluations for mortality.
> Body Weights: Body weights were obtained prior to exposure on day 0 and on post-exposure days 1, 3, 7, 10 and 14.
- Necropsy of survivors performed: Yes. Animals were euthanised by Isoflurane® anaesthesia following by exsanguination. The major organ systems of the cranial, thoracic and abdominal cavities were examined for all animals.

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.5 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

None of the animals died during exposure or the 14-day observation period.

Clinical signs:

other: Immediately following exposure, gasping was observed for one male and bilateral ptosis (partial closure of the eye) was noted for four males and five females. There were no other toxicologically significant clinical observations. Wet tan material on the e

Body weight:

Slight body weight losses (2 to 7 g) were noted for four males and three females from day 0 to 1. All animals surpassed their initial (day 0) body weight by day 3 and were considered normal.

Gross pathology:

There were no gross findings at the scheduled necropsy.

Table 1: Mean body weights (g)

Sex	Pre-exposure	Day 1	Day 3	Day 7	Day 10	Day 14
Male	159	156	171	193	203	221
Female	146	145	152	155	157	163

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the study, the LC50 of the test material was found to be greater than 5.5 mg/L when male and female rats were exposed to the test material as a dust aerosol for four hours.

Executive summary:

The acute inhalation toxicity of the test material was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 403, EPA OPPTS 870.1300, EU Method B.2 and JMAFF Nousan 4200.

During the study test material was administered to a single group of five male and five female albino rats, over a period of 4 hours, via nose-only exposure as a dust aerosol at a concentration of 5.5 mg/L. The exposure aerosol was characterised by a mass median aerodynamic diameter of 2.5 µm with a geometric standard deviation of 2.45. Following exposure, all animals were maintained for a 14-day observation period. Parameters evaluated were mortality, clinical observations, body weights and gross necropsy.

None of the animals died during exposure or the 14-day observation period. Gasping was observed for one male and bilateral ptosis was noted for nine animals (four males, five females) immediately following exposure. There were no other toxicologically significant clinical observations immediately following exposure. Dried red material around the nose, ptosis or complete closure of the eyes and/or yellow material on the urogenital area were noted for one female during the first week. There were no other toxicologically significant clinical observations during the 14-day observation period. Nine of the 10 animals had slight body weight losses from day 0 to 1. All animals surpassed their initial (day 0) body weight by day 3 and were considered normal. There were no gross findings at the scheduled necropsy.

Under the conditions of the study, the LC50 of the test material was found to be greater than 5.5 mg/L when male and female rats were exposed to the test article as a dust aerosol for four hours.

 

Reference 2

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1300 (Acute inhalation toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.2 (Acute Toxicity (Inhalation))

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF 2-1-3

Deviations:

no

GLP compliance:

yes

Test type:

traditional method

Limit test:

yes

Specific details on test material used for the study:

TSN314667
Batch: ENBK-169021-016
Purity: 95.9%

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Animal Breeding Facility, Jai Research Foundation
- Females (if applicable) nulliparous and non-pregnant: [yes/no] yes
- Age at study initiation: 8 to 9 weeks
- Weight at study initiation: (g) Minimum: 252.5, Maximum: 258.1 for male and Minimum: 197.2, Maximum: 205.2 for female
- Fasting period before study: no
- Housing: Polypropylene rat cages covered with stainless steel grid tops were used. Autoclaved clean rice husk was used as the bedding material. Wooden blocks were used as enrichment material. Two to three rats/cage.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 23 °C
- Humidity (%): 56 to 65%
- Air changes (per hr): Minimum 15 air changes/hour
- Photoperiod (hrs dark / hrs light): 12 h artificial light and 12 h darkness, light hours being 06:00 - 18:00 h, maintained through automatic timer.

Route of administration:

other: inhalation: dust/aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

3.42 µm

Geometric standard deviation (GSD):

1.64

Details on inhalation exposure:

The 4-hour acute inhalation exposure was carried out using nose-only inhalation exposure equipment supplied by CH Technologies, USA. This dynamic inhalation exposure system consists of a flow-past nose-only inhalation exposure chamber with Perspex rat exposure/restraint tubes, air compressor, standard positive pressure control unit (flow meters, pressure gauge, vacuum lines) rotating brush generator (dry powder), cascade impactor, temperature and humidity probe (thermo-hygrometer), oxygen monitor, carbon dioxide monitor, and open face gravimetric filter sampler.

The rats were exposed using a nose-only inhalation exposure system with a dynamic air flow rate of 727 to 728 air changes per hour and the air inflow rate was maintained at 25 litres per minute, ensuring an adequate oxygen content of at least 19% and a homogeneous, evenly distributed, respirable test aerosol with a mass median aerodynamic diameter (MMAD) particle size range of 1-4 μm with a geometric standard deviation (GSD) in the range of 1.5 to 3.0. The exposure atmosphere carbon dioxide level was less than 1%.

The dynamic inhalation equipment has 2 main parts namely outer plenum and inner plenum. The inner plenum receives the test item which is delivered to each exposure tube. The outer plenum is the exhaust side. The excess air along with the test item flows into the outer plenum. The inner plenum volume is 862 cm3 and the outer plenum volume is 1,200 cm3 so the total volume is 2062 cm3 or 2.062 Liters. The exposure unit (outer plenum) is made of stainless steel with 12 portholes to accommodate transparent Perspex rat exposure tubes. These exposure tubes are accommodated in the portholes of the inhalation chamber. The adjustable unit of the exposure tube is set in such a way that rats breathe the test item aerosol through the window panel of the exposure tube. Observations during the inhalation experiment will be made through transparent Perspex rat exposure tubes. The outlet unit is connected to a suction pump. The outgoing air from the chamber passes through an impinger containing 1.0% sodium hydroxide solutions and moisture traps.

The dust generator system is manufactured by PALAS GmbH and supplied by CH Technologies (USA), Inc. The dust generator system is a long assembly of tubing having several tube fittings attached. The feed system consists of a feed stock reservoir, and the generating system consists of brush housing with integrated brush, spacer holder, up/down schalter, inlet of air for dispersion, connection of compressed air with reducer, and switch for manual or external controls (air, brush speed, schalter speed).

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

5.49 mg/L

No. of animals per sex per dose:

5/gender

Control animals:

no

Details on study design:

All rats were observed for any signs of toxicity and mortality at hourly intervals during the 4-hour exposure period and at 1 and 2 hours after the exposure on the day of exposure. Subsequently, the surviving rats were observed twice a day for morbidity and mortality for a period of 14 days following exposure. The clinical signs were recorded once a day.

Body weights were recorded prior to exposure on day 0 and on post exposure days 1, 3, 7 and 14.

At the end of the observation period, all rats were sacrificed by intraperitoneal administration of thiopentone sodium and subjected to gross pathological examination. This consisted of an external examination and the opening of the nasal passage, abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded.

Statistics:

As this study was conducted as a limit study at the time-weighted average (TWA) exposure concentration of 5.49 mg/L air, no statistical analysis was required to calculate the LC50.

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.49 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

No mortality was observed in rats exposed for 4 hours to an aerosol concentration of 5.49 mg aminopyralid TGAI/L air (TWA).

Clinical signs:

other: No treatment related clinical signs were observed in any of the rats exposed to an aerosol concentration of 5.49 mg aminopyralid TGAI/L air.

Body weight:

A decrease in mean body weight was observed in both the sexes of rats at day 1 post exposure, with respect to day 0 mean body weight (before exposure). From day 3 body weight was recovered and continued increasing on days 7 and 14 with respect to day 0 mean body weight (before exposure).

Gross pathology:

External examination of terminally sacrificed rats did not reveal any abnormality. Visceral examination of terminally sacrificed rats did not reveal any abnormality.

Interpretation of results:

GHS criteria not met

Conclusions:

4-hr inhalation LC50 (male/female rats) > 5.49 mg/L

Executive summary:

The acute inhalation toxicity of aminopyralid TGAI was investigated in the Wistar rats (RCCHan:WIST). A single set of 5 male and 5 female rats (approximately 8-9 weeks old and weighing 197.2-258.1 g on the day of exposure, were exposed to dust aerosol of aminopyralid TGAI with a time-weighted average concentration of 5.49 mg/L air using a nose-only inhalation exposure system. The mass median aerodynamic diameter (MMAD) of aminopyralid TGAI aerosol was determined to be 3.42 µm with a geometric standard deviation (GSD) of 1.64. The rats were exposed for 4 hours followed by a 14-day post exposure observation period during which body weight and clinical observations were recorded. All animals were sacrificed and a necropsy was performed.

 

No treatment related mortality or clinical signs were observed.

 

A decrease in mean body weight was observed in both the sexes of rats at day 1 post exposure, with respect to day 0 mean body weight (before exposure). From day 3 body weight was recovered and continued increasing on days 7 and 14 with respect to day 0 mean body weight (before exposure). 

 

No internal or external abnormalities were recorded at the post-mortem examination.

 

In conclusion, based on the study results, the 4-hour acute inhalation median lethal concentration (LC₅₀value) of aminopyralid TGAI inWistar rats (male and female combined) was found to be greater than5.49 mgaminopyralid TGAI/L air.

 

Based on the results of this study, an indication of the classification for aminopyralid TGAI is as follows:

Globally Harmonized System of Classification and Labelling of Chemicals (GHS 2017): Unclassified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LC50

Value:

5.49 mg/m³ air

Quality of whole database:

Two studies were conducted under GLP conditions and in accordance with standardised guidelines; the studies were therefore assigned a reliability score of 1 in line with the criteria of Klimisch et al. (1997). The overall quality of the database is good.

Acute toxicity: via dermal route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 June 2001 to 30 August 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1200 (Acute Dermal Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF Acute Dermal Toxicity Study, 2000

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

Purity: 94.5%

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: ca. 8 weeks
- Weight at study initiation: 122 - 176 g
- Housing: animals were housed two or three per cage in stainless steel cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 1 week

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 3 °C
- Humidity: 40 - 70 % (relative)
- Air changes: 12 - 15 air changes per hour
- Photoperiod: 12 hours of darkness / 12 hours of light (06:00 to 18:00)

Type of coverage:

semiocclusive

Vehicle:

other: the test material was moistened with 0.5 % aqueous methylcellulose

Details on dermal exposure:

The dorsal side of the trunk of each rat was clipped free of fur the day prior to application of the test material. The test material, moistened with 0.3 mL of 0.5 % aqueous methylcellulose, was administered to the back of the rats (approximately 10 % of the surface area of the rats) and covered with a 6 x 6 cm gauze/cotton patch. The gauze patch was held in place by elastic tape and wrap. The wrappings were removed after approximately a 24-hour exposure period and observations were recorded for irritation at the application site. The skin was wiped thoroughly with a water-moistened soft disposable towel and dried with a soft disposable towel.

Duration of exposure:

24 hours

Doses:

5000 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: A Detailed Clinical Observation (DCO) was conducted for all rats prior to test material administration for comparison with the observations recorded throughout the study. Animals were observed a minimum of two times on the day of treatment. A DCO was done each day (including weekends and holidays) during the study. Hand-held and open-field observations included a careful physical examination. The rats were weighed pre-study, the day of treatment and on days 2, 8, and 15.
- Necropsy of survivors performed: Yes, necropsy was performed on all animals. Animals were anaesthetised by inhalation of carbon dioxide and euthanised by decapitation after clamping of the trachea.
- Other examinations performed: The eyes were examined in situ using a moistened glass microscope slide applied to the corneal surface. Following inspection of the externum and body orifices, the nasal, cranial, oral, thoracic, and abdominal cavities were opened and the visceral organs were examined both in situ and following dissection, and tissues were not saved.

Statistics:

Means and standard deviations were calculated for body weights. The data were evaluated for statistical outliers by a sequential test (Grubbs, 1969); however, outliers were not routinely excluded from statistical analysis.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Mortality:

All animals survived the two-week observation period.

Clinical signs:

other: Clinical observations consisted of perioral soiling in one male and two females on test day 1 or 2 and periocular soiling in one male on test day 1. There was also an abrasion or scratches observed on the dermal test site of one male and one female rat fr

Gross pathology:

There were no gross pathologic observations.

Table 1: Mean body weights (g)

	Days on test
Sex	-1	1	2	8	15
Male	168.9	173.2	162.8	200.8	227.3
Female	127.1	128.6	118.0	137.6	146.6

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this study, the acute dermal LD50 of the test material in male and female Fischer 344 rats was greater than 5000 mg/kg.

Executive summary:

The acute dermal toxicity of the test material was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 402, EU Method B.3, EPA OPPTS 870.1200 and JMAFF (2000).

During the study, five male and five female Fischer 344 rats received a single 24-hour exposure of 5000 mg/kg body weight of test material moistened with 0.5 % aqueous methylcellulose. Parameters evaluated during the two-week observation period included body weights, detailed clinical observations, and gross pathological changes.

All animals survived the two-week observation period. Clinical observations consisted of perioral soiling in one male and two females on test day 1 or 2 and periocular soiling in one male on test day 1. There was also an abrasion or scratches observed on the dermal test site of one male and one female rat from test day’s 2-5 or test day 4, respectively. The test material dried and adhered to these two animals, thus causing the abrasion/scratches during test material removal. All animals lost weight by test day 2, but gained weight over the remainder of the study. There were no gross pathologic observations.

Therefore, under the conditions of this study, the acute dermal LD50 of test material in male and female Fischer 344 rats was greater than 5000 mg/kg bw.

Reference 2

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

TSN314667
Batch: ENBK-169021-016
Purity: 95.9%

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Animal Breeding Facility, Jai Research Foundation
- Females (if applicable) nulliparous and non-pregnant: [yes/no] yes
- Age at study initiation: 11-13 weeks
- Weight at study initiation: (g) Minimum: 218.1, Maximum: 227.2
- Fasting period before study: no
- Housing: Polypropylene rat cages covered with stainless steel grid tops were used. Autoclaved clean rice husk was used as the bedding material. Wooden blo ck was provided as enrichment material.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 to 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 23 °C
- Humidity (%): 49 to 66%
- Air changes (per hr): Minimum 15 air changes/hour
- Photoperiod (hrs dark / hrs light): 12 h artificial light and 12 h darkness, light hours being 06:00 - 18:00 h

Type of coverage:

semiocclusive

Vehicle:

other: distlled water

Details on dermal exposure:

The required amount (1090.5 to 1136.0 mg) of aminopyralid TGAI (moistened with 0.5 mL distilled water) was applied over the treatment area (approximately 7 cm × 5 cm body surface area, previously clipped), and kept in place using porous gauze dressing and a non-irritating tape (semi-occlusive) for a 24-hour exposure period.

Duration of exposure:

24 hours

Doses:

5000 mg/kg

No. of animals per sex per dose:

3

Control animals:

no

Details on study design:

The rats were observed for signs of toxicity and mortality within 0.5 and at 2, 4 and 6 hours post-dermal application on the day of dosing (day 0). Subsequently, the rats were observed twice a day for morbidity and mortality for a period of 14 days following dermal application. Erythema and oedema were recorded at 24, 48 and 72 hours post patch removal in all three rats. The clinical signs were recorded once a day. Individual body weight was recorded prior to dermal application on day 0 and on days 7 and 14 post-dermal application.

At the end of 14 days observation period, all rats were euthanised by carbon dioxide asphyxiation. They were subjected to a gross pathological examination consisting of an external and internal examination and opening of abdominal and thoracic cavities.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality was observed in rats treated with 5000 mg aminopyralid TGAI/kg body weight.

Clinical signs:

other: No treatment related clinical signs were observed in any of the rats treated with 5000 mg aminopyralid TGAI/kg body weight. No erythema and oedema were observed at 24, 48 and 72 hours post patch removal in all three rats.

Gross pathology:

External examination of terminally sacrificed female rats did not reveal any abnormalities of pathological significance.

Visceral examination of female rats sacrificed at termination did not reveal any lesions.

Interpretation of results:

GHS criteria not met

Conclusions:

Acute Dermal LD50 (female rats) > 5000 mg/kg

Executive summary:

In an acute dermal toxicity study, three female Wistar rats(approximately 11-13 weeks old and weighing 218.1-227.2g at dosing)were dermally exposed toaminopyralid TGAIfor 24 hoursat the dose of 5000 mg/kg body weight). The required amount (1090.5 to 1136.0 mg) ofaminopyralid TGAI(moistened with 0.5 mL distilled water) was applied over the treatment area (approximately 7 cm × 5 cm body surface area, previously clipped), and kept in place using porous gauze dressing and a non-irritating tape (semi-occlusive) for a 24-hour exposure period. Initially only one rat was treated. Based on the observations at 24 hours post patch removal, two additional rats were dosed sequentially. At the end of the 24-hour exposure period, the residual test item was removed with cotton soaked in distilled water. The skin reactions of each rat were observed at 24, 48 and 72 hours post patch removal. The rats were observed for a period of 14 days (in which mortality, clinical signs and body weights were recorded) after which rats were sacrificed and subjected to gross necropsy examination.

 

No mortality or clinical signs were observed in any animal throughout the study period.

 

No erythema or oedema were evident at 24, 48 and 72 hours post patch removal in any of the three rats.

 

No internal or external abnormalities were observed at the post-mortem examination.

 

Based on the results of this study, the acute dermal median lethal dose (LD₅₀value) of aminopyralid TGAI in female Wistar rats was found to be greater than 5000 mg/kg body weight.

 

Based on the results of this study, an indication of the classificationforaminopyralid TGAIisas follows:

 

Globally Harmonized System of Classification and

Labelling of Chemicals (GHS 2017)                                          : Unclassified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

5 000 mg/kg bw

Quality of whole database:

Two studies were conducted under GLP conditions and in accordance with standardised guidelines; the studies were therefore assigned a reliability score of 1 in line with the criteria of Klimisch et al. (1997). The overall quality of the database is good.

Additional information

Oral

The acute oral toxicity of the test material was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 401, EU Method B.1, EPA OPPTS 870.1100 and JMAFF (2000).

During the study, five male and five female Fischer 344 rats received test material at a dose of 5000 mg/kg body weight as a 50 % mixture in 0.5 % aqueous methylcellulose by single dose gavage. Parameters evaluated during the two-week observation period included body weights, detailed clinical observations, and gross pathological changes.

Under the conditions of this study, the acute oral LD50 of the test material in male and female Fischer 344 rats was greater than 5000 mg/kg. A second study conducted under guideline OECD 423, EU Method B.1, EPA OPPTS 870.1100 and JMAFF 2 -1 -1 with female Wistar rats. In this study, the acute oral LD50 was also found to be greater than 5000 mg/kg.

Inhalation

The acute inhalation toxicity of the test material was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 403, EPA OPPTS 870.1300, EU Method B.2 and JMAFF Nousan 4200. During the study test material was administered to a single group of five male and five female albino rats, over a period of 4 hours, via nose-only exposure as a dust aerosol at a concentration of 5.5 mg/L. The exposure aerosol was characterised by a mass median aerodynamic diameter of 2.5 µm with a geometric standard deviation of 2.45. Following exposure, all animals were maintained for a 14-day observation period. Parameters evaluated were mortality, clinical observations, body weights and gross necropsy.

None of the animals died during exposure or the 14-day observation period. Gasping was observed for one male and bilateral ptosis was noted for nine animals (four males, five females) immediately following exposure. There were no other toxicologically significant clinical observations immediately following exposure. Dried red material around the nose, ptosis or complete closure of the eyes and/or yellow material on the urogenital area were noted for one female during the first week. There were no other toxicologically significant clinical observations during the 14-day observation period. Nine of the 10 animals had slight body weight losses from day 0 to 1. All animals surpassed their initial (day 0) body weight by day 3 and were considered normal. There were no gross findings at the scheduled necropsy.

Under the conditions of the study, the LC50 of the test material was found to be greater than 5.5 mg/L when male and female rats were exposed to the test article as a dust aerosol for four hours.  

A second study was conducted in male and female Wistar rats in accordance with standardised guidelines OECD 403, EPA OPPTS 870.1300, EU Method B.2 and JMAFF 2 -1 -3. This second study confirmed that the 4 -hour LC50 was greater than5.49 mg/L.

Dermal

The acute dermal toxicity of the test material was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 402, EU Method B.3, EPA OPPTS 870.1200 and JMAFF (2000).

During the study, five male and five female Fischer 344 rats received a single 24-hour exposure of 5000 mg/kg body weight of test material moistened with 0.5 % aqueous methylcellulose. Parameters evaluated during the two-week observation period included body weights, detailed clinical observations, and gross pathological changes.

All animals survived the two-week observation period. Clinical observations consisted of perioral soiling in one male and two females on test day 1 or 2 and periocular soiling in one male on test day 1. There was also an abrasion or scratches observed on the dermal test site of one male and one female rat from test day’s 2-5 or test day 4, respectively. The test material dried and adhered to these two animals, thus causing the abrasion/scratches during test material removal. All animals lost weight by test day 2, but gained weight over the remainder of the study. There were no gross pathologic observations.

Therefore, under the conditions of this study, the acute dermal LD50 of test material in male and female Fischer 344 rats was greater than 5000 mg/kg bw.

A second study was conducted according to OECD guideline 402 with female Wistar rats. The acute dermal LD50 was also greater than 5000 mg/kg in this study.

Justification for classification or non-classification

In accordance with the criteria for classification as defined in Annex I, Regulation (EC) No. 1272/2008, the substance does not require classification with respect to acute toxicity.

In accordance with the criteria for classification as defined in Annex VI, Directive 67/548/EEC (DSD), the substance does not require classification with respect to acute toxicity.