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Toxicity to birds

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long-term toxicity to birds: reproduction test
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23 May - 16 Nov 1990
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
according to guideline
EPA OPP 71-4 (Avian Reproduction Test)
Version / remarks:
Data Requirement FIFRA 163.71-4
Temperature stability analysis delayed
GLP compliance:
Dose method:
Analytical monitoring:
Details on preparation and analysis of diet:
The test compound was dissolved in corn oil and acetone then placed in a separatory funnel and added to the feed while mixing for 10 minutes in a 30-quart bowl of a Hobart Mixer. Acetone was also used as a rinsing agent for the glassware used in the preparation of the diets. Fresh batches of diet were prepared weekly and stored in the freezer until used. At the end of each test week all uneaten diet was destroyed by incineration and fresh feed was offered to the birds.

Homogeneity studies conducted both during the pilot study and the definitive study showed NTN 33893 was mixed homogeneously in the gamebird ration at concentrations between 30 and 240 ppm. The coefficient of variation for the analysis during the definitive study was 8.3 and 1.33% respectively for the low and high concentrations. Results from the room temperature stability analysis showed NTN 33893 was stable in the avian ration at room temperature. The recoveries were between 91 and 115% with a relative standard deviation of 6.40 %. The linearity of instrument response is good between 1- and 7-ppm standard concentrations.

The mean of the measured concentrations determined from five time periods were 35.7, 60.7, 126, and 243 ppm (119, 101, 105 and 101 % nominal) for the 30-, 60-, 120-, and 240-ppm nominal concentrations respectively. NTN 33893 was not found in the Control samples.

At the beginning of the pilot study, acetonitrile was used as the extraction solvent. The solvent was changed to methanol since it has better extractability with NTN 33893 for room temperature stability study samples.

Test organisms (species):
Colinus virginianus
Details on test organisms:
Pen-reared bobwhite quail (Colinus virginianus) were apparently healthy and phenotypically indistinguishable from wild birds. All birds were from the same hatch and were approximately 20 weeks of age at test initiation. The birds were approaching their first breeding season and had not been used in previous testing. Test birds were acclimated to the laboratory facilities for 28 days prior to initiation of the test. Mortality during the acclimation period was 2% of the birds received for the study (5 of 266). Necropsy observations of these birds revealed dehydration and emaciation due to the birds inability to adjust to the caging.
Limit test:
Total exposure duration (if not single dose):
20 wk
No. of animals per sex per dose and/or stage:
Each dose contained 18 pairs of birds with one male and one female per pen.
Control animals:
yes, concurrent no treatment
Nominal and measured doses / concentrations:
Nominal: 30, 60, 120 and 240 mg of technical NTN 33893 as active ingredient per kg of diet.
Measured: 35.7, 60.7, 126 and 243 mg a.s./kg of diet
Details on test conditions:
Adult birds were housed indoors in pens manufactured by Shore Manufacturing that measured approximately 55L X 27W X 24H cm with sloping floors. Pens were constructed of stainless steel wire grid and stainless steel sheeting. A group of three cages shared a common feeder
and water trough. Feeders were designed to minimize spillage. Each week, fresh feed was placed into clean feeders for each group of mated pairs and presented to the birds. Waterers were changed and water added as necessary to provide a constant supply of clean potable water. Cage bedding was changed weekly. Birds were placed into clean cages once every three weeks.
Adult birds were all housed in a single room with a temperature range during the course of the study of 68 ± 4°F and a relative humidity range of 40 to 70%. The photoperiod in the adult study room was maintained by a Modicon Light Control System. The photoperiod for the first seven weeks of the study was 7 hours of light per day. At the beginning of week eight it was then increased to 17 hours of light per day until the sacrifice of the adult birds. A 40 minute "dawn/dusk" cycle was used in light/dark transition. The birds received a minimum of six foot-candles of illumination through the study.
Hatchlings were placed in batteries of brooding pens (90L X 72W X 24H cm). The external walls and ceilings of each pen were constructed of galvanized wire mesh and galvanized sheeting. The floors were of galvanized wire mesh. Cage bedding (compressed alfalfa pellets) was changed at least once each week. Thermostats in the brooding compartment of each pen were set to maintain a temperature of approximately 100°F from the time of hatching through day 14 post-hatch. The photoperiod for the hatchlings was maintained by time clock at 12 hours light per day.
Details on examinations and observations:
All adult birds were observed at least once daily throughout the study for signs of toxicity or abnormal behavior.
Individual adult body weights were recorded at study initiation, then weekly through week 8, and at terminal sacrifice.
Feed consumption was recorded, by feeder, weekly throughout the study.
Details on reproductive parameters:
Eggs were collected twice daily through the week and once each day on weekends and holidays. Once each week eggs were removed from the egg cooler and candled for shell cracks.
Eggs were candled again on day 11 of incubation to determine embryo viability (fertility) and on day 21 to determine embryo survival.
Every other week of the egg laying period, eggs were collected from each laying hen for eggshell strength and thickness measurements.
Reference substance (positive control):
not specified
Key result
Duration (if not single dose):
20 wk
Dose descriptor:
Effect level:
126 mg/kg diet
Conc. / dose based on:
act. ingr.
Basis for effect:
body weight
Mortality and sub-lethal effects:
No compound-related mortality occurred during the study.
There were no compound-related symptoms of toxicity noted during the study.
Based on the type of observations recorded and on the number of birds with notable lesions, all observations gross necropsy were considered incidental and not related to the test material.
There were no significant differences in female body weight between the control and treated groups. The mean body weight of the 243 ppm male birds was significantly less than controls at the end of the adult exposure.
There was no statistical difference between control and treated groups in feed consumption.
Effects on reproduction:
There were no statistically significant differences between the control treated groups in any reproductive parameter.
There was no significant difference in survival of the offspring at any treatment level.
The overall mean hatchling body weight was significantly less than control for all concentrations, however, there were only sporadic differences with no clear dose response trend for lower body weights at any one concentration.
This difference in offspring body weight is not biologically significant since there was no significant decrease in hatch percentage, no significant difference in the percent survival and no clear dose response trend in the weekly hatchling or survival body weight. In fact a weight gain for the offspring was equal to or greater than the control for each group.

Please refer to "overall remark/ attached background material" field for result tables.

Validity criteria fulfilled:
not specified
The present guideline study was conducted in compliance with GLP. Under the test conditions used, the overall NOEC (20 wk) for Colinus virginianus was estimated to be 126 mg/kg diet based on effects on male body weight.

Description of key information

The overall NOEC (20 weeks) for Colinus virginianus was estimated to be 126 mg a.s./kg diet.

Key value for chemical safety assessment

Long-term EC10, LC10 or NOEC for birds:
126 mg/kg food

Additional information

A key study (1991) investigating the chronic effects of test material on bird species following EPA OPP Guideline 71-4, under GLP was available. The birds were exposed to the nominal dietary concentrations of 30, 60, 120 and 240 mg a.s./kg feed, alongside with a control. Mean measured concentrations from week 1 to week 20 after application ranged from 101% to 119% of nominal concentrations. Based on the mean measured concentrations after 20 weeks of exposure to the test material, no effects on the reproductive performance of Colinus virginianus were reported up to the third concentration tested (NOEC (20wk) = 126 mg a.s./kg diet).


A supportive study was conducted for 20 weeks according to the EPA OPP Guideline 71-4 and GLP with Anas platyrhynchos resulting in NOEC (20wk) of 128 mg a.s./kg diet, based on the reproductive parameters.

The acute dietary toxicity of test material to bird species has been investigated in three supportives studies. The studies were conducted with standard species (Anas platyrhynchos and Colinus virginianus) according to EPA OPP Guidelines 71-2, and OECD 205, under GLP conditions. The results from the 5 days-dietary tests resulted in LC50 values ranging between of 392 to > 4797 mg a.i./kg diet.


Additionally, birds' mortality was evaluated in three acute oral tests, resulting in a LD50 ranging from 31 to 283 mg a.s./kg body weight according to EPA OPP Guidelines 71-1.