ethyl 5,5-diphenyl-2-isoxazoline-3-carboxylate

Toxicity to birds

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to birds: dietary toxicity test

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

20 - 28 May 1997

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 205 (Avian Dietary Toxicity Test)

Version / remarks:

adopted in 1984

Deviations:

yes

Remarks:

Acclimatisation period only 3 days, instead of 7 days recommended; 17h light instead of 12-16h recommended

Qualifier:

according to guideline

Guideline:

EPA OPP 71-2 (Avian Dietary Toxicity Test)

Version / remarks:

adopted in 1982

GLP compliance:

yes

Dose method:

feed

Analytical monitoring:

yes

Details on preparation and analysis of diet:

DIET PREPARATION
- Description and nutrient analysis of basal diet provided in study report: yes
- Preparation of doses: A 5 kg final mixture of each of the 5 dietary concentrations of the test substance was prepared and stored deep frozen.

HOMOGENEITY AND STABILITY OF TEST MATERIAL IN DIET
- How often was homogeneity and stability tested: Directly after preparation 3 samples from each of the lowest and highest concentrations were thawed and analysed for achieved test substance content, homogeneity and, after 11 days storage at room temperature, for stability.
- Results of homogeneity analysis (range of values):
- Nominal concentration (mg/kg feed): 0, 312.5 and 5000
- Concentration analysed (mg/kg feed): not detectable, 241.0 - 254.4 and 4071 - 4112
- % of nominal (mean): -, 80 and 82
- Mixing procedure adequate and variance between nominal and actual dosage acceptable (yes/no): yes
- Results of stability analysis (range of values):
- Nominal concentration (mg/kg feed): 0, 312.5 and 5000
- Concentration after 11 days storage at room temperature (mg/kg feed): not detectable, 260.3 - 302.1 and 4776 - 5112
- % of nominal (mean): -, 90 and 100

Test organisms (species):

Anas platyrhynchos

Details on test organisms:

TEST ORGANISM
- Common name: mallard duck
- Source: Zuchtgeflugelhof B. Franzsander, Schöneck-Oberdorfelden, Federal Republic of Germany, Batch No. EK 3/97
- Age at test initiation (mean and range, SD): 10 days
- Weight at test initiation (mean and range, SD): mean = 105.4 g, s = ± 14.81 g, n = 70

Limit test:

no

Total exposure duration (if not single dose):

5 d

Remarks:

followed by a 3-day recovery period with untreated diet

No. of animals per sex per dose and/or stage:

10 animals per dose (sex not reported)

Control animals:

yes

Nominal and measured doses / concentrations:

Nominal: 0, 312.5, 625, 1250, 2500 or 5000 ppm
For measured concentrations see "Any other information on results incl. tables" below

Details on test conditions:

ACCLIMATION
- Acclimation period: 3 days
- Feeding: The ducklings received a special duckling starter ration, SSNIFF® V 6310. Food and water were available ad libitum.
- Health (any disease or mortality observed): no mortality occurred in the study population due to health or any other unknown factor

PEN SIZE AND CONSTRUCTION MATERIALS
- Description: rearing and maintenance batteries 81 cm long, 78 cm wide and 22 cm high
- Floor covering: wire-mesh floors
- Caging: group

NO. OF BIRDS PER REPLICATE
- For negative control: 10
- For treated: 10

NO. OF REPLICATES PER GROUP
- For negative control: 1
- For treated: 1

TEST CONDITIONS (range, mean, SD as applicable)
- Temperature: 28 - 32 °C
- Room temperature: 20 - 23 °C
- Relative humidity (%): ca. 30 - 60
- Photoperiod: 17 hours light and 7 hours darkness

RANGE FINDING STUDY
- Results used to determine the conditions for the definitive study: The dose levels selected for this study were based on the results of the acute oral toxicity study in the Mallard duck which yielded a LD50 value of greater 2000 mg/kg body-weight.

Details on examinations and observations:

MORTALITY / CLINICAL SIGNS
- Time schedule for examinations: clinical signs and the mortality rate were recorded twice daily on work-days and once daily on weekends and public holidays

BODY WEIGHT
- Time schedule for examinations: at the start of acclimatisation (day -3), at the start of treatment (day 1), the end of treatment (day 6) and at the end of the 3-day recovery period at the end of study (day 9)

FOOD CONSUMPTION (if feeding study)
- Time schedule for examinations: recorded for each group for days 1-6 (5-day treatment period) and 6-9 (3-day recovery period)

PATHOLOGY
- Dose groups that were examined: all groups including controls

Duration (if not single dose):

5 d

Dose descriptor:

LC50

Effect level:

> 5 000 other: ppm (diet)

Conc. / dose based on:

test mat.

Basis for effect:

mortality

Duration (if not single dose):

5 d

Dose descriptor:

LC50

Effect level:

> 1 675 mg/kg bw/day

Conc. / dose based on:

test mat.

Basis for effect:

mortality

Duration (if not single dose):

5 d

Dose descriptor:

NOEL

Effect level:

5 000 other: ppm (diet)

Conc. / dose based on:

test mat.

Basis for effect:

other: mortality, clinical signs, body weight, food consumption, autopsy findings

Mortality and sub-lethal effects:

MORTALITY
No mortality occurred in any dose group.

CLINICAL SIGNS
- Results: No clinical signs of toxicity occurred in any dose group at any time during the study up to and including the 5000 ppm dietary level.

BODY WEIGHT AND FOOD CONSUMPTION
- Results: Food consumption and body weight gain remained unaffected by the test substance up to and including the 5000 ppm dietary level. During the 3-day recovery period both, food consumption and body weight gain showed no difference compared with the control groups.

PATHOLOGY
- Results: Dissection of the ducklings killed at the end of the study revealed no macroscopically visible abnormalities.

Table 1: Concentration control at start of treatment (after 11 days storage in a deep-freezer)

Nominal concentration (mg/kg)	Sample	Concentration found (mg/kg)	%of nominal	Mean value % of nominal
0	A	n.d.	-	-
	B	n.d.	-
	C	n.d.	-
312.5	A	293.0	94	90
	B	294.3	94
	C	256.3	82
625	A	625	100	98
	B	590	94
	C	616	98
1250	A	1168	93	93
	B	1152	92
	C	1158	93
2500	A	2258	90	94
	B	2337	93
	C	2447	98
5000	A	4724	94	94
	B	4746	95
	C	4654	93

 

Table 2: Validity criteria for OECD 205.

Criterion from the guideline	Outcome	Validity criterion fulfilled
Mortality in controls should not exceed 10%.	0%	yes
The concentraion of the substance being tested has been satisfactorily been maintained in the diet (Should be at least 80% of nominal concentration) throughout the first five days of the test period.	11 days after mixture preparation: 90 - 98%	yes
The lowest treatment level should not result in compound-related mortality	No compoud-related effects were detected at the lowest test concentration.	yes

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Reference 2

Endpoint:

short-term toxicity to birds: acute oral toxicity test

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

03 - 17 Dec 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPP 71-1 (Avian Acute Oral Toxicity Test)

GLP compliance:

yes

Dose method:

gavage

Analytical monitoring:

yes

Vehicle:

yes

Details on preparation and analysis of diet:

DIET PREPARATION
- Description and nutrient analysis of basal diet provided in study report: yes
- Preparation of doses: The test substance was prepared at the stated concentration in a homogeneous aqueous solution of methylcellulose (1 % w/v) by means of a magnetic stirrer.
- Type and function of solvent/vehicle: methylcellulose (1%) in deion. water, tylose

HOMOGENEITY AND STABILITY OF TEST MATERIAL IN DIET
- When and at what dose levels were samples of treated food analyzed for stability and concentration during the study: 3 mg/mL after 0 and 4 h; 24 mg/mL and 200 mg/mL after 4 h
- Results of homogeneity and stability analysis (range of values): see "Any other information on results, incl. tables" below
- Mixing procedure adequate and variance between nominal and actual dosage acceptable (yes/no): yes

Test organisms (species):

Colinus virginianus

Details on test organisms:

TEST ORGANISM
- Common name: Bobwhite Quail
- Source: Hoechst AG - Hoechst Marion Roussel Preclinical Development - Drug Safety
- Age at test initiation (mean and range, SD): approx. 5 months
- Weight at test initiation (mean and range, SD): males: 190 g (177 g - 210 g); females 179 g (158 g - 207 g)
- Sexes used: male and female

Limit test:

yes

Remarks:

single dose

Post exposure observation period:

up to 14 days

No. of animals per sex per dose and/or stage:

5 males and 5 females per dose

Control animals:

yes

Nominal and measured doses / concentrations:

Nominal: 0 and 2000 mg/kg b.w.

Details on test conditions:

ACCLIMATION
- Acclimation period: at least two weeks
- Feeding: A special diet prepared by SSNIFF® GmbH and tap water in drinking troughs were given. Food and water were available ad libitum.
- Fasting period before study: 15 hours before dosing and 2 hours afterwards

FEED WITHHOLDING PERIOD BEFORE DOSING
- No. of days: 15 hours before dosing and 2 hours afterwards

PEN SIZE AND CONSTRUCTION MATERIALS
- Description: rearing and maintenance batteries, 81 cm long, 78 cm wide and 22 cm high
- Floor covering: wire-mesh floors
- Caging: group

NO. OF BIRDS PER REPLICATE
- For vehicle control: 10
- For treated: 10

NO. OF REPLICATES PER GROUP
- For vehicle control: 1
- For treated: 1

TEST CONDITIONS (range, mean, SD as applicable)
- Temperature: about 20°C
- Relative humidity (%): 20 - 50 %
- Photoperiod: 8 hours light and 16 hours darkness

RANGE FINDING STUDY
- Results used to determine the conditions for the definitive study: The dose selection for this study was based on the acute toxicity in rats which is > 2000 mg/kg body weight.

Details on examinations and observations:

The birds were weighed individually on days 1, 4, 8 and 15 of the 15-day observation period. The total food consumption per group was also recorded for days 1 - 4, 4 - 8 and 8-1 5 after treatment (day 1 = day of treatment). At the end of the observation period, the birds were killed by carbon dioxide asphyxiation, dissected and examined both externally and internally (i.e. examination of internal body cavity contents) for macroscopically visible changes.

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Conc. / dose based on:

test mat.

Basis for effect:

mortality

Remarks on result:

other: single dose

Mortality and sub-lethal effects:

MORTALITY
No mortality occurred in the treated or control groups.

CLINICAL SIGNS
- Results: No clinical signs of toxicity occurred in the treated dose groups.

BODY WEIGHT AND FOOD CONSUMPTION
- Results: Body weight and food consumption were not affected by the test substance.

PATHOLOGY
- Results: Necropsy revealed no morphological changes.

Table 1: Homogeneity and stability

Nominal conc. mg/mL	Nominal conc. %	Sample	Found in mg/mL after 0h	Found in mg/mL after 4h	% of nominal after 0h	% of nominal after 4h
3	0,3	A	2.85	2.60	95	87
		B	2.75	2.84	92	95
		C	2.65	2.78	88	93
24	2,4	A	-	20.7	-	86
		B	-	19.9	-	83
		C	-	20.0	-	83
200	20	A	-	197	-	99
		B	-	208	-	104
		C	-	202	-	101

Table 2: Validity criteria for OECD 213

Criterion from the guideline	Outcome	Validity criterion fulfilled
The average mortality for the total number of controls must not exceed 10% at the end of the test.	< 10%	yes
The LD50 of the toxic standard meets the specified range (0.10 – 0.35 µg a.i./bee, 24 h).	Not specified	Not applicable

Validity criteria fulfilled:

not applicable

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Reference 3

Endpoint:

short-term toxicity to birds: acute oral toxicity test

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

10 - 24 Dec 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPP 71-1 (Avian Acute Oral Toxicity Test)

Version / remarks:

adopted in 1982

GLP compliance:

yes

Dose method:

gavage

Analytical monitoring:

yes

Vehicle:

yes

Details on preparation and analysis of diet:

DIET PREPARATION
- Description and nutrient analysis of basal diet provided in study report: yes
- Preparation of doses: The test substance was prepared at the stated concentration in a homogeneous aqueous solution of methylcellulose (1 % w/v) by means of a magnetic stirrer.
- Type and function of solvent/vehicle: methylcellulose (1%) in deion. water, tylose

HOMOGENEITY AND STABILITY OF TEST MATERIAL IN DIET
- When and at what dose levels were samples of treated food analyzed for stability and concentration during the study: 3 mg/mL after 0 and 4 h; 24 mg/mL and 200 mg/mL after 4 h
- Results of homogeneity analysis (range of values): see "Any other information on results, incl. tables" below
- Mixing procedure adequate and variance between nominal and actual dosage acceptable (yes/no): yes

Test organisms (species):

Anas platyrhynchos

Details on test organisms:

TEST ORGANISM
- Common name: Mallard duck
- Source: Geflugelhof B. Franzsander Im Brinkort 25, Delbruck-Hagen, Germany
- Age at test initiation (mean and range, SD): approx. 4 months
- Weight at test initiation (mean and range, SD): males: 1281 g (1103 g - 1510 g); females: 1136 g (1042 g - 1272 g)
- Sexes used: male and female

Limit test:

yes

Remarks:

single dose

Post exposure observation period:

up to 14 days

No. of animals per sex per dose and/or stage:

5 males and 5 females per dose

Control animals:

yes

Nominal and measured doses / concentrations:

Nominal: 0 and 2000 mg/kg bw.

Details on test conditions:

ACCLIMATION
- Acclimation period: two weeks
- Feeding: A special diet prepared by SSNIFF® GmbH and tap water in drinking troughs were given. Food and water were available ad libitum.
- Fasting period before study: 15 hours before dosing and 2 hours afterwards

FEED WITHHOLDING PERIOD BEFORE DOSING
- No. of days: 15 hours before dosing and 2 hours afterwards

PEN SIZE AND CONSTRUCTION MATERIALS
- Description: pens (180 x 300 cm)
- Floor covering: smooth concrete floors
- Caging: group

NO. OF BIRDS PER REPLICATE
- For vehicle control: 5
- For treated: 5

NO. OF REPLICATES PER GROUP
- For vehicle control: 1
- For treated: 1

TEST CONDITIONS (range, mean, SD as applicable)
- Temperature: 20°C
- Relative humidity (%): 30 - 60
- Photoperiod: 8 hours light and 16 hours darkness

RANGE FINDING STUDY
- Results used to determine the conditions for the definitive study: The dose selection for this study was based on the acute toxicity in rats which is
1740 (1251 -2245) mg/kg body weight

Details on examinations and observations:

After treatment, any signs of toxicity, mortality rate and time of death were recorded. The birds were weighed individually on days 1,4, 8 and 15 of the 15-day observation period. The total food consumption per group was also recorded for days 1 - 4, 4 - 8 and 8-1 5 after treatment (day 1 = day of treatment). At the end of the observation period, the surviving birds (incl. controls) were killed by intravenous injection of T61®, dissected and examined both externally and internally (i.e. examination of internal body cavity contents) for macroscopically visible changes.

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Conc. / dose based on:

test mat.

Basis for effect:

mortality

Remarks on result:

other: single dose

Mortality and sub-lethal effects:

MORTALITY
No mortality occurred in the treated or control groups.

CLINICAL SIGNS
- Results: No clinical signs of toxicity occurred in the treated dose groups.

BODY WEIGHT AND FOOD CONSUMPTION
- Results: Body weight and food consumption were not affected by the test substance

PATHOLOGY
- Results: Necropsy revealed no morphological changes.

Table 1: Validity criteria for OECD 213

Criterion from the guideline	Outcome	Validity criterion fulfilled
The average mortality for the total number of controls must not exceed 10% at the end of the test.	< 10%	yes
The LD50 of the toxic standard meets the specified range (0.10 – 0.35 µg a.i./bee, 24 h).	Not specified	Not applicable

Validity criteria fulfilled:

not applicable

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Reference 4

Endpoint:

short-term toxicity to birds: dietary toxicity test

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

05 - 13 Jun 1997

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 205 (Avian Dietary Toxicity Test)

Version / remarks:

adopted in 1984

Deviations:

yes

Remarks:

Acclimatisation period only 3 days, instead of 7 days recommended; 17h light instead of 12-16h recommended

Qualifier:

according to guideline

Guideline:

EPA OPP 71-2 (Avian Dietary Toxicity Test)

Version / remarks:

adopted in 1982

GLP compliance:

yes

Dose method:

feed

Analytical monitoring:

yes

Details on preparation and analysis of diet:

DIET PREPARATION
- Description and nutrient analysis of basal diet provided in study report: yes
- Preparation of doses: A 3 kg final mixture of each of the 5 dietary concentrations of the test substance was prepared and stored deep frozen.

HOMOGENEITY AND STABILITY OF TEST MATERIAL IN DIET
- How often was homogeneity and stability tested: Directly after preparation 3 samples from each of the lowest and highest concentrations were taken and analysed for achieved test substance content, homogeneity and, after 44 days storage at room temperature, for stability
- Results of homogeneity analysis (range of values):
- Nominal concentration (mg/kg feed): 0, 312.5 and 5000
- Concentration analysed (mg/kg feed): not detectable, 275.6 - 298.2 and 4461 - 5047
- % of nominal: -, 91 and 95
- Mixing procedure adequate and variance between nominal and actual dosage acceptable (yes/no): yes
- Results of stability analysis (range of values):
- Nominal concentration (mg/kg feed): 0, 312.5 and 5000
- Concentration after 11 days storage at room temperature (mg/kg feed): not detectable, 285.7 - 290.4 and 4577 - 4883
- % of nominal: -, 92 and 94

Test organisms (species):

Colinus virginianus

Details on test organisms:

TEST ORGANISM
- Common name: Bobwhite quail
- Source: Hoechst Marion Roussel Deutschland GmbH, Global Preclinical Development - Drug Safety, Batch No. KVW 2/97
- Age at test initiation (mean and range, SD): 10 days
- Weight at test initiation (mean and range, SD): mean = 29.3 g, s = ± 0.97 g, n = 7 test groups

Limit test:

no

Total exposure duration (if not single dose):

5 d

Remarks:

followed by a 3-day recovery period with untreated diet

No. of animals per sex per dose and/or stage:

10 animals per dose

Control animals:

yes

Nominal and measured doses / concentrations:

Nominal: 0, 312.5, 625, 1250, 2500 and 5000 ppm
For measured concentrations see "Any other information on results incl. tables" below

Details on test conditions:

ACCLIMATION
- Acclimation period: 3 days
- Feeding: The chicks received a special avian complete rearing diet, SSNIFF® V 6310. Food and water were available ad libitum.
- Health (any disease or mortality observed): no mortalities observed

PEN SIZE AND CONSTRUCTION MATERIALS
- Description: rearing and maintenance plastic boxes (floor space approx. 4200 cm2 / group)
- Floor covering: strewn with quartz sand with a wire-mesh overlay
- Caging: group

NO. OF BIRDS PER REPLICATE
- For negative control: 10
- For treated: 10

NO. OF REPLICATES PER GROUP
- For negative control: 1
- For treated: 1

TEST CONDITIONS (range, mean, SD as applicable)
- Temperature: about 32°C
- Room temperature: 19 - 22 °C
- Relative humidity (%): 40 - 70
- Photoperiod: 17 hours light and 7 hours darkness

RANGE FINDING STUDY
- Results used to determine the conditions for the definitive study: The dose levels selected for this study were based on the results of the acute oral toxicity in the Bobwhite quail which is greater 2000 mg/kg bodyweight.

Details on examinations and observations:

MORTALITY / CLINICAL SIGNS
- Time schedule for examinations: recorded twice daily on work days and once daily on weekends and public holidays

BODY WEIGHT
- Time schedule for examinations: The birds were weighed by group at the start of acclimatisation (day -3), at the start of treatment (day 1), the end of treatment (day 6) and at the end of the 3-day recovery period at the end of study (day 9).

FOOD CONSUMPTION (if feeding study)
- Time schedule for examinations: recorded for days 1-6 (5-day treatment period) and 6-9 (3-day recovery period).

PATHOLOGY
- Dose groups that were examined: all groups including controls

Duration (if not single dose):

5 d

Dose descriptor:

LC50

Effect level:

> 5 000 other: ppm (diet)

Conc. / dose based on:

test mat.

Basis for effect:

mortality

Duration (if not single dose):

5 d

Dose descriptor:

LC50

Effect level:

> 980 mg/kg bw

Conc. / dose based on:

test mat.

Basis for effect:

mortality

Duration (if not single dose):

5 d

Dose descriptor:

NOEL

Effect level:

5 000 other: ppm (diet)

Conc. / dose based on:

test mat.

Basis for effect:

other: mortality, clinical signs, body weight, food consumption, autopsy findings

Duration (if not single dose):

5 d

Dose descriptor:

NOEL

Effect level:

980 mg/kg bw

Conc. / dose based on:

test mat.

Basis for effect:

other: mortality, clinical signs, body weight, food consumption, autopsy findings

Mortality and sub-lethal effects:

MORTALITY
- No mortality attributable to the treatment with the test substance occurred in any dose group.
- One chick in the 625 ppm group died on the last day of treatment. In the absence of clinical signs and mortalities in the higher dose groups, this death was considered to be incidental and not related to the test substance.

CLINICAL SIGNS
- Results: No clinical signs of toxicity occurred in any dose group at any time during the study up to and including the 5000 ppm dietary level.

BODY WEIGHT AND FOOD CONSUMPTION
- Results: Food consumption and body weight gain remained unaffected by the test substance up to and including the 5000 ppm dietary level. During the 3-day recovery period, both food consumption and body weight gain showed no difference compared with the control groups.

PATHOLOGY
- Results: Dissection of the surviving chicks killed at the end of the study and of the chick which died in the 625 ppm group revealed no macroscopically visible morphological abnormalities.

 Concentration control at start of treatment (after 11 days storage in a deep-freezer)

Nominal concentration (mg/kg)	Sample	Concentration found (mg/kg)	%of nominal	Mean value % of nominal
0	A	n.d.	-	-
	B	n.d.	-
	C	n.d.	-
312.5	A	264.3	85	87
	B	281.1	90
	C	268.7	89
625	A	612	98	94
	B	566	91
	C	585	94
1250	A	1163	93	97
	B	1251	100
	C	1207	97
2500	A	2672	107	101
	B	2509	100
	C	2367	95
5000	A	4839	97	107
	B	5215	104
	C	5959	119

 

Table 2: Validity criteria for OECD 205.

Criterion from the guideline	Outcome	Validity criterion fulfilled
Mortality in controls should not exceed 10%.	0%	yes
The concentraion of the substance being tested has been satisfactorily been maintained in the diet (Should be at least 80% of nominal concentration) throughout the first five days of the test period.	44 days after mixture preparation: 87 - 107%	yes
The lowest treatment level should not result in compound-related mortality	No compoud-related effects were detected at the lowest test concentration.	yes

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Reference 5

Endpoint:

long-term toxicity to birds: reproduction test

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 206 (Avian Reproduction Test)

Version / remarks:

adopted in 1984

Qualifier:

according to guideline

Guideline:

other: Pesticide Assessment Guidelines. FIFRA Subdivision E. Hazard Evaluation: Wildlife and Aquatic Organisms. Subsection 71-4. Environmental Protection Agency, Office of Pesticide Programs, October 1982

GLP compliance:

yes

Dose method:

feed

Analytical monitoring:

yes

Details on preparation and analysis of diet:

DIET PREPARATION
- Description and nutrient analysis of basal diet provided in study report: yes
- Preparation of doses: For each concentration, test diets were prepared weekly from a premix containing the test substance mixed with a small quantity of basal diet. Each required nominal dietary concentration of the test substance was prepared from a portion of the premix by further addition of basal diet. When needed during the test, additional diet was prepared. Premixes were prepared as needed, generally every three to four weeks. The nominal dietary concentrations were not adjusted for the purity of the test substance.

HOMOGENEITY AND STABILITY OF TEST MATERIAL IN DIET
- How often was homogeneity and stability tested: Homogeneity of the test substance in the diet was evaluated by collecting three samples from each treatment group on Day 0 of Week 1. Diet samples were collected from the control group and each treatment group after being held for one, two, four, eight and 15 days at room temperature to verify the amount of the test substance under actual test conditions.
- Results of homogeneity and stability analysis:
- Homogeneity was shown to be satisfactory in the week 1 mix at 40, 200 and 1000 ppm, i.e. mean values obtained for top, middle and bottom samples were within the acceptable range 85 - 110% of nominal and these mean % values differed by < 10%.
- Stability was shown to be satisfactory over the time of use of the diet i.e. % nominal values declined by < 11% over 8 days storage at room temperature.
(See "Any other information about results incl. tables" below)

Test organisms (species):

Anas platyrhynchos

Details on test organisms:

TEST ORGANISM
- Common name: mallard duck
- Source: Whistling Wings, 113 Washington Street, Hanover, Illinois 61041, USA
- Age at test initiation (mean and range, SD): 24 weeks
- Weight at test initiation (mean and range, SD): 870 g - 1357 g
- Sexes used: male and female
- Disease free: apparently healthy

Limit test:

no

Total exposure duration (if not single dose):

21 wk

No. of animals per sex per dose and/or stage:

16 males and 16 females

Control animals:

yes

Nominal and measured doses / concentrations:

Nominal: 0, 40, 200 and 1000 ppm
Measured: see "Any other information about results incl. tables" below

Details on test conditions:

ACCLIMATION
- Acclimation period: 7 weeks
- Feeding: Untreated basal diet available ad libitum

PEN SIZE AND CONSTRUCTION MATERIALS
- Description: batteries of pens measuring approximately 75 x 90 x 45 cm high
- Floor covering: galvanized wire grid
- Caging: pairs (male and female)

NO. OF BIRDS PER REPLICATE
- For negative control: 2
- For treated: 2

NO. OF REPLICATES PER GROUP
- For negative control: 16
- For treated: 16

TEST CONDITIONS (range, mean, SD as applicable)
- Temperature: 21.8 °C ± 1.5 °C
- Relative humidity (%): 61 ± 21 (SD)
- Photoperiod: 8 h or less light for the first 10 weeks; 17 h light from week 11
- Ventilation: The air handling system in the study room was designed to vent up to 15 room air volumes every hour and replace it with fresh air.

RANGE FINDING STUDY
In a six-week dietary reproduction range-finding study conducted previously at these laboratories, there were no treatment related mortalities at any of the test concentrations (500, 1000 and 5000 ppm). Additionally, there were no overt signs of toxicity, or effects upon body weight, feed consumption or reproductive condition.

Details on examinations and observations:

All adult birds were observed at least once daily throughout the study for signs of toxicity or abnormal behavior.

MORTALITY / CLINICAL SIGNS
A record was maintained for all mortalities and observations. All birds that died during the study were necropsied. As soon as practical after the death of the bird, the penmate was sacrificed and necropsied.

BODY WEIGHT
Birds were weighed individually at the start of the treatment period (Day 0 of Week 1), on Weeks 2, 4, 6 and 8 and at adult termination. Body weights were not measured during egg laying because of the adverse effects handling may have had on egg production.

FOOD CONSUMPTION (if feeding study)
Feed consumption was measured weekly for each pen and is presented on a weekly basis. Feed consumption was determined by weighing the freshly filled feeder when presented to the birds, recording the amount of any additional diet added during the week, and weighing the feeder and remaining feed at the end of the feeding period. No attempt was made to quantify the amount of feed wasted by the birds, as the wasted feed is normally scattered and mixed with water and excreta.

PATHOLOGY
At necropsy, records of the external appearance and macroscopic abnormalities of tissues and organs of the thoracic and abdominal cavities were made for adult birds dying or euthanized during the course of the test and for those surviving to termination.

Details on reproductive parameters:

Eggs and Embryos:
Records were maintained for each pen and each week of the numbers of eggs that were laid, cracked, abnormal, set and hatched and the numbers of viable and live three week embryos. Egg shell thickness was recorded for each pen and week as appropriate

Offspring:
Records were kept of the numbers of hatchlings and offspring surviving for 14 days (14-day survivors) per pen, per week. Offspring body weights were determined at hatching and at 14 days of age for surviving ducklings.

Duration (if not single dose):

21 wk

Dose descriptor:

NOEC

Effect level:

200 other: ppm in diet

Conc. / dose based on:

test mat.

Basis for effect:

reproductive parameters

Duration (if not single dose):

21 wk

Dose descriptor:

NOEC

Effect level:

22.4 mg/kg bw/day

Conc. / dose based on:

test mat.

Basis for effect:

reproductive parameters

Mortality and sub-lethal effects:

MORTALITY
- Results: No treatment related mortalities occurred during the test.
- Remarks: While no mortalities occurred in the control or 200 ppm treatment groups, one incidental mortality occurred in both the 40 and the 1000 ppm treatment groups. Due to the nature of the lesions observed at necropsy, the mortalities were considered not to be treatment related.

CLINICAL SIGNS
- Results: No clinical signs of toxicity were seen at any of the concentrations tested.

BODY WEIGHT
- Results: There were no treatment related effects upon adult body weight at any of the concentrations tested, and any differences from the control group in body weight were not statistically significant at any of the body weight intervals.

FOOD CONSUMPTION (if feeding study)
- Results: There were no treatment related effects upon feed consumption at the 40, 200 or 1000 ppm test concentrations.

PATHOLOGY
- Results: All findings were considered incidental to treatment.

Effects on reproduction:

Egg Production, Embryo Viability, Hatching and Offspring Survival:
There were no treatment related effects upon reproductive performance at the 40 or 200 ppm treatment groups, and any differences from the control group were not statistically significant for any of the reproductive parameters measured. However, at the 1000 ppm test concentration there was a reduction in overall reproductive performance that was considered to be treatment related. Effects upon reproductive performance in the 1000 ppm treatment group were
apparent in both egg production and hatchability values (eggs laid/maximum laid and hatchlings/live 3-week embryos). The effects upon egg production and hatchability also were reflected as reductions in the overall number of hatchlings and 14-day old survivors (hatchlings/eggs set, hatchlings/maximum set, 14-day old survivors/eggs set and 14-day old survivors/maximum set). While not statistically significant, differences from the control means for the affected parameters were considered to be treatment related.

Egg Shell Thickness:
There were no treatment-related effects upon egg shell thickness at the 40, 200 or 1000 ppm test concentrations, and any differences from the control group were not statistically significant.

Table 1: Homogeneity and stability: Homogeneity was tested by analysing top, middle and bottom samples.

Study week	% nominal
	40 ppm	200 ppm	1000 ppm
Week 1 – top	89.6	88.7	94.3
Week 1 – middle	85.4	89.2	93.7
Week 1 – bottom	85.2	92.3	95.1
Week 1 stability day 0	84.3	89.3	89.9
Week 1 stability day 8	77.0	79.6	87.7
Week 1 stability day 15	76.0	77.0	85.2
Week 4	87.7	96.2	102.3
Week 8	96.4	101.2	97.7
Week 12	102.1	113.5	107.4
Week 21	92.1	90.9	94.1

Table 2: Validity criteria for OECD 206.

Criterion from the guideline	Outcome	Validity criterion fulfilled
Mortality in controls should not exceed 10%.	0%	yes
Average number of 14 -day-old survivors per hen in the controls should be at least 14 for mallard duck.	31	yes
The average eggshell thickness for the control group should be at least 0.34 mm for mallard duck.	0.378 mm	yes
If the recommended concentration scheme is followed and no effect on reproduction is detected, the results may be reported as the NOEC being greater than the highest concentration tested.	not applicable	yes

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Description of key information

LC50 (5 d): > 5000 ppm (diet) / > 1675 mg/kg/day (Anas platyrhynchos, OECD 205)

NOEC (21 wk): 200 ppm in diet/22.4 mg/kg/day (Anas platyrhynchos, OECD 206)

Additional information

Information on the toxicity to birds is not required under Regulation (EC) No 1907/2006, but to complete the data set additional information is provided.

A total of GLP five studies is available investigating the toxicity of the substance to birds, following the prevailing guidelines OECD 205/206 and/or US EPA OPP 71-1 and 2.

In a 5-day study with mallard ducks (Anas platyrhynchos) the LC50 was found to be above 5000 ppm in diet or 1675 mg/kg body weight and day. A single-dose study with a 14-day observation period reports the LD50 to be above 2000 mg/kg body weight. As the result of a 5-day dietary study with bobwhite quails (Colinus virginianus), an LC50 > 5000 ppm in diet or > 980 mg/kg body weight is reported with the quails exhibiting no behavioural or clinical/pathological abnormalities that can be ascribed to the test substance . In a single-dose test with a 14-day observation period employing bobwhite quails (Colinus virginianus) exposed to 2000 mg test substance/kg body weight, no mortality occurred.

In summary, it is concluded that the LD50 for oral toxicity to birds is greater than 5000 ppm in diet or 1675 mg/kg body weight.

A 21-week dietary reproduction study with mallard ducks (Anas platyrhynchos) revealed a NOEC of 200 ppm in diet or 22.4 mg/kg body weight and day.