cyproconazole (ISO); (2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-1-(1H-1,2,4-triazol-1-yl)butan-2-ol

Toxicity to terrestrial arthropods

Link to relevant study record(s)

Reference

Endpoint:

toxicity to bees: acute oral

Remarks:

and contact test

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not reported

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline followed

Principles of method if other than guideline:

Contact test: Mortality was recorded at 24 hours after dermal application.
Oral test: Organisms were fed a honey-water solution containing the test substance for 24 hours. Mortality was recorded at 24 hours after application.

GLP compliance:

not specified

Application method:

other: Contact and Oral

Analytical monitoring:

no

Vehicle:

yes

Remarks:

acetone (contact exposure)

Details on preparation and application of test substrate:

- Contact toxicity: To apply the test substance, the bees were anaesthetised with carbon dioxide during 15 - 30 seconds. Then, the insects were laid, ventral surface up, on filter paper in a Petri dish. 1.0 microliter drops of the test substance, dissolved in analytical grade acetone, were then placed on one of the metatarsi with a microapplicator.

- Oral toxicity: 0.6 mL of a mixture, containing the appropriate concentration of the test substance and 67% honey in water. was fed to each group of bees, i.e. 20 microliters per bee. Each concentration was administered to two lots of 30 bees.

Test organisms (species):

Apis mellifera

Animal group:

Hymenoptera (honeybees)

Details on test organisms:

TEST ORGANISM
- Common name: Worker honeybee
- Source: The test facility's own colonies, located in Witterswil, near Basle

Study type:

laboratory study

Limit test:

no

Total exposure duration:

24 h

Test temperature:

- Contact toxicity test: 25 °C
- Oral toxicity test: 24 ± 2 ˚C

Humidity:

- Contact toxicity test: 65% relative humidity
- Oral toxicity test: 50 - 60% relative humidity

Details on test conditions:

TEST SYSTEM (Contact toxicity)
- Test container: 200 mL polystyrene cup
- Cover: With a wire mesh cover
- No. of organisms per container: 10
- No. of replicates per treatment group: 3
- No. of replicates per negative control: 3

OTHER TEST CONDITIONS (Contact toxicity)
- Photoperiod: 16 hours light and 8 hours darkness
- Humidity: 65 % relative humidity

EFFECT PARAMETERS MEASURED (Contact toxicity)
24 hours after treatment, surviving and killed bees were counted. Mortality was expressed in percent, and corrected for natural mortality.

TEST SYSTEM (Oral toxicity)
- Test container: styropor-boxes (28.5 x 25 x 10.5 cm) with glass front and windows sealed with wire mesh in the side and back walls for aeration.
- No. of organisms per container: 30
- No. of replicates per treatment group:2
- No. of replicates per negative control: 2

OTHER TEST CONDITIONS (Oral toxicity)
When the bees had taken all the test substance solution ( 0.5 to 3 hours) , they were given unlimited water and food paste (sugar+ honey, 3:1). It was assumed that the 0.6 mL of the test substance solution would be redistributed by the bees between themselves so that they would receive equal doses.
- Photoperiod: Daylight supplemented by fluorescent tubes disposed 20 cm above the cages during 10 hours per day.
- Humidity: 50 - 60% relative humidity

EFFECT PARAMETERS MEASURED (Oral toxicity)
24 hours after the fungicide administration, surviving and killed insects were counted.

Nominal and measured concentrations:

- Nominal concentration (Contact): 0 (negative control) and 100 µg a.i./bee
- Nominal concentration (Oral): 0 (negative control), 40, 200 and 1000 µg a.i./bee

Reference substance (positive control):

yes

Remarks:

Parathion - ethyl

Key result

Duration:

24 h

Dose descriptor:

LD50

Effect conc.:

> 100 µg per animal

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: Contact exposure

Key result

Duration:

24 h

Dose descriptor:

LD50

Effect conc.:

> 1 000 µg per animal

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: Oral exposure

Details on results:

An overview of the results is provided in Table 1 - Table 3 in 'Any other information on results incl. tables'.
At 1000 µg ai/bee 17% mortality was observed via oral exposure. No other clinical observations reported. No contact mortality was reported at highest dose level of 100 µg ai/bee over 24 hour exposure. No other signs of toxicity were reported.

Results with reference substance (positive control):

Via contact exposure, the LD50 of the reference substance was determined to be 0.14 µg a.i./bee via contact exposure. While via oral exposure, the LD50 was determinded to be 0.13 µg a.i./bee.

Reported statistics and error estimates:

LD50 values were calculated by linear regression analysis, using the logit method. Since two assays were carried out for each fungicide, the resulting two LD50 values were pooled by computing a weighted geometric mean according to Finney (1971).

Table 1. Acute Contact Activity

compound	µg a.i. per bee	rep. no.	number of bees			% mortality
			Total	Alive	Dead
Test substance 95.2% a.i.	100 100 100	1 2 3	10 10 10	10 10 10	0 0 0	0
Untreated check	- - -	1 2 3	10 10 10	10 10 10	0 0 0	0

  

Table 2. Oral Activity

compound	µg a.i. per bee	rep. no.	number of bees			% mortality
			Total	Alive	Dead
Test substance 95.2% a.i.	1000 1000 200 200 40 40	1 2 1 2 1 2	30 30 30 30 30 30	23 27 30 27 30 30	7 3 0 3 0 0	17   5   0
Untreated check	- -	1 2	30 30	30 30	0 0	0

 Remarks:

μg a.i. per bee: amount of active ingredient, applied to the bees, in micrograms

rep.no. : number of repetition

number of bees, total: used bees per repetition

number of bees, alive/dead: number of bees per repetition alive or dead, 24 hours after treatment

% mortality: mortality in percent

Table 3. Results of acute contact (topical application) and oral toxicities to worker honeybees

Compound	Contact		Oral
	LD 50	SE	LD50	SE
Test substance   Parathion-ethyl	>100   0.14	-   0.01	> 1000   0.13	-   0. 01

Note. LD 50 and standard errors (SE) expressed asµg a.i./bee, 24 hours after application.

rep.no. : number of repetition

number of bees, total: used bees per repetition

number of bees, alive/dead: number of bees per repetition alive or dead, 24 hours after treatment

% mortality: mortality in percent

 

Validity criteria fulfilled:

yes

Conclusions:

Based on the findings, the 24-hour LD50 of the test substance in honey bee was > 1000 μg ai/bee via the oral route and >100 μg ai/bee via contact exposure route.

Executive summary:

The acute contact and oral toxicities to terrestrial arthropods were investigated using honeybee (Apis mellifera) as test organism. The study did not follow guideline and was not in compliance with GLP. However, the study meets generally accepted scientific principles and the test procedures and results are well documented, so that the study is considered as reliable (with restrictions). For oral test, two replicates of 30 bees per group were fed a honey-water solution containing the test substance at nominal levels of 40, 200 and 1000 µg ai/bee. When the bees had taken all the of the treated feed solution (0.5 - 3 hours), untreated diet was offered ad libitum. A control group fed with an untreated honey-water solution was also added. Temperature and humidity were maintained at 24 ± 2 ˚C and 50 - 60 % relative humidity, respectively. Mortality was recorded at 24 hours after application. Results are based on nominal concentrations per bee. 

For contact test, three replicates of 10 bees per group were exposed to the test substance administered topically in a 1 µL-droplet to the thorax of each bee, at a nominal level of 100 µg ai/bee (limit test) for 24 hours. Acetone was used to prepare the solutions. One additional group was included as untreated control. Temperature and humidity were maintained at 25 °C and 65 % relative humidity, respectively. Mortality was recorded at 24 hours after application. Results are based on nominal concentrations per bee. 

The results indicate that at 1000 µg ai/bee treatment, 17 % mortality was observed via oral exposure, and no other clinical observations were reported. No contact mortality was reported at the limit dose level of 100 µg ai/bee over 24-hour exposure. No other signs of toxicity were reported. Based on the findings, the 24-hour LD50 of the test substance in honeybee was > 1000 µg ai/bee via the oral route and >100 µg ai/bee via contact exposure route.

Description of key information

24-h LD50 > 100 µg a.i./bee (contact dosing), Apis mellifera, mortality, no guideline followed, Donat 1985

24-h LD50 > 1000 µg a.i./bee (oral dosing), Apis mellifera, mortality, no guideline followed, Donat 1985

Additional information

The acute contact and oral toxicities to terrestrial arthropods were investigated using honeybee (Apis mellifera) as test organism. The study did not follow guideline and was not in compliance with GLP. However, the study meets generally accepted scientific principles and the test procedures and results are well documented, so that the study is considered as reliable (with restrictions). For oral test, two replicates of 30 bees per group were fed a honey-water solution containing the test substance at nominal levels of 40, 200 and 1000 µg ai/bee. When the bees had taken all the of the treated feed solution (0.5 - 3 hours), untreated diet was offered ad libitum. A control group fed with an untreated honey-water solution was also added. Temperature and humidity were maintained at 24 ± 2 ˚C and 50 - 60 % relative humidity, respectively. Mortality was recorded at 24 hours after application. Results are based on nominal concentrations per bee. 

For contact test, three replicates of 10 bees per group were exposed to the test substance administered topically in a 1 µL-droplet to the thorax of each bee, at a nominal level of 100 µg ai/bee (limit test) for 24 hours. Acetone was used to prepare the solutions. One additional group was included as untreated control. Temperature and humidity were maintained at 25 °C and 65 % relative humidity, respectively. Mortality was recorded at 24 hours after application. Results are based on nominal concentrations per bee. 

The results indicate that at 1000 µg ai/bee treatment, 17 % mortality was observed via oral exposure, and no other clinical observations were reported. No contact mortality was reported at the limit dose level of 100 µg ai/bee over 24-hour exposure. No other signs of toxicity were reported. Based on the findings, the 24-hour LD50 of the test substance in honeybee was > 1000 µg ai/bee via the oral route and >100 µg ai/bee via contact exposure route.