spirodiclofen (ISO);3-(2,4-dichlorophenyl)-2-oxo-1-oxaspiro[4.5]dec-3-en-4-yl 2,2-dimethylbutyrate

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

This toxicity study is classified as acceptable and satisfies the guideline requirements for the aquatic nonvascular plant toxicity study with the green alga, Selenastrum capricornutum.

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Version / remarks:

15 Nov. 1989

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: EPA-Guideline 540/9-86- 134 (Environmental Protection Agency, Hazard Evaluation Division, Standard Evaluation Procedure - Non-Target Plants Growth and Reproduction of Aquatic Plants, Tiers 1 and 2

Version / remarks:

June, 1986

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

Revised Version No. L 383
A179 (29 Dec 1992).

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

7 June 1984

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: GOP Standards

Version / remarks:

(OECD, C (81)
30 (Final), May 12, 1981

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

96-hour acute toxicity study, the cultures of the green alga, Pseudokirchneriella subcapitata, were exposed to spirodiclofen technical at the nominal concentrations of 20, 40 and 60 µg/L. The study included a control. The measured concentrations were 20.0, 41.5 and 61.6 µg/L. The test was a static design.

Parameters measured including the growth inhibition/other toxicity symptoms - Inhibition was measured by calculating growth rate and biomass.
Measurement technique for cell density and other end points - Photometric technique to estimate cell numbers.

Vehicle:

yes

Details on test solutions:

-150-mL aliquots of test solution in 300-mL Erlenmeyer flasks.
-Details of growth medium: Details of nutrient solution are included in report. The media was referenced to an EEC directive (1992); ISO 8692 (1989); and OECD 201 (1984). Chelator: Na2EDTA x 2 H2O (100 µg/L) Carbon source: Na HCO3 pH at initiation = 8.3 pH at termination = 9.7
Deionized water was used to prepare nutrient media. pH = 8.3 at test initiation and 9.7 at termination.
-Controls: The control solution is not specified as nutrient media alone or a solvent control. Since there is only one control in the design it is most likely a solvent control. 6 replicates for the control and 6 at each treatment level
-Chemical name of vehicle (organic solvent, emulsifier or dispersant): acidified methanol (2.5% glacial acetic acid). The concentration of solvent in the treatment solutions is not specified in the report.
-Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): Used a stock solution of Spirodiclofen technical dissolved in acidified methanol (2.5% glacial acetic acid)

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

Name: (class and species) green alga, Pseudokirchneriella subcapitata
Strain: Strain 61.81, Collection of Algal Cultures, Inst. Plant Physiology, Univ.
Gottingen, Germany.
Source: laboratory stock culture
Age of inoculum: cultures were 3 days old at test initiation.
Method of cultivation: Sterile algal cultures were grown in nutrient media and
renewed once per week.

Acclimation: Not reported.
The pre-culture which was used to inoculate the study was grown in the same media that was used in the test solutions. The environmental conditions for testing and culturing were the same. Cells were examined microscopically to look for malformations. No abnormalities were reported.

Test type:

static

Water media type:

freshwater

Total exposure duration:

96 h

Test temperature:

Temperature = 23.5 ± 1.5­­ °C

Nominal and measured concentrations:

Nominal concentrations: 60, 40 and 20 µg/L.
Initial measured on Day 0: 61.6, 41.5 and 19.8 µg/L

Details on test conditions:

- Incubation facility: Environmental chamber
- Test vessel: Glass flasks sealed with cotton wool plugs. 150-mL aliquots of test solution in 300-mL Erlenmeyer flasks:
- Aeration: 3 revolutions per minute. No aeration.
- Initial cells density: 1E+4 cells/mL
- Control end cells density: Controls started at 1E+4 and ended at 301E+4 cells/ml.
- No. of organisms per vessel: The control solution is not specified as nutrient media alone or a solvent control. Since there is only one control in the design it is most likely a solvent control.
6 replicates for the control and 6 at each treatment level

Growth Medium
- Standard medium used: yes
- Detailed composition if non-standard medium was used: Details of nutrient solution are included in report. The media was referenced to an EEC directive (1992); ISO 8692 (1989); and OECD 201 (1984). Chelator: Na2EDTA x 2 H2O (100 µg/L) Carbon source: Na HCO3 pH at initiation = 8.3, pH at termination = 9.7.

Dilution Water:
- Source/preparation of dilution water: Deionized water was used to prepare nutrient media.
- pH: pH = 8.3 at test initiation and 9.7 at termination
- Photoperiod, Light intensity and quality: Lighting was continuous (24 hour light phase) at 6945 lux.

Observation:
- Parameters measured including the growth inhibition/other toxicity symptoms
- Inhibition was measured by calculating growth rate and biomass.
- Measurement technique for cell density and other end points
- Photometric technique to estimate cell numbers.
- Observation intervals: every 24 hours
- Water quality was acceptable: Yes
- Were raw data included: Yes

- Range finding study: no range find results were mentioned in the report
- Test concentrations: 60, 40 and 20 µg/L.
- Results used to determine the conditions for the definitive study:

Reference substance (positive control):

yes

Key result

Duration:

96 h

Dose descriptor:

LOEC

Effect conc.:

> 60 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

>= 60 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Key result

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

> 60 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 60 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

act. ingr.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 60 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

act. ingr.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 60 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

act. ingr.

Basis for effect:

biomass

Details on results:

There were no adverse effects on growth or cell morphology during this 96 hour exposure.

Results with reference substance (positive control):

K2Cr2O7 test concentrations were 0.10, 0.18, 0.32, 0.56, 1.0 and 1.8 mg/L. 72 hr EC50 was 0.78 mg/L and 1.57 mg/L for biomass and growth rate, respectively.

Reported statistics and error estimates:

The EC50 for growth of biomass (EbC50) and for algal growth rate (ErC50) were calculated using probit analyses and the slopes of the regression lines were calculated following Litchfield and Wilcoxon. The NOEC and LOEC were calculated by an analysis of variance (Dunnett's-Test).

Table 3: Effect of spirodiclofen technical on algal growth (green alga, Pseudokirchneriella subcapitata)

-	Cell density (1E+4)			96 hours
-					% inhibition
Treatment nominal (µg/L)	24 hours	48 hours	72 hours	Cell count	biomass	Growth rate
Solvent control	6.94	35.2	139.45	301.44	0	0
20	7.10	37.7	147.90	317.58	-5.8	-0.9
40	7.41	39.0	150.87	322.56	-7.8	-1.2
60	8.2	40.1	156.76	325.68	-10.5	-1.4

Validity criteria fulfilled:

yes

Conclusions:

The study is acceptable. The EC50 is greater than 60 µg ai/L and the NOEC is 60 µg ai/L. Within the limits of solubility, the compound does not inhibit algal growth.

Executive summary:

In a 96-hour acute toxicity study, the cultures of the green alga, Pseudokirchneriella subcapitata, were exposed to spirodiclofen technical at the nominal concentrations of 20, 40 and 60 µg/L. The study included a control. The measured concentrations were 19.8, 41.5 and 61.6 µg/L. The test was a static design. The NOEC was 60 µg/L. The 96-hour EC50 based on growth rate and biomass was greater than 60 µg/L. This was the highest level tested since the spirodiclofen is insoluble at higher concentrations. There was no inhibition in the treatment levels as compared to the controls for growth rate or biomass.
No abnormalities were noted. This toxicity study is classified as acceptable and satisfies the guideline requirements for the aquatic nonvascular plant toxicity study with the green alga, Selenastrum capricornutum.
Results Synopsis
Test Organism: green alga (Pseudokirchneriella subcapitata or Selenastrum capricornutum)
Test Type: Static
96-hr EC50 >60 µg/L
96-hr NOEC >=60 µg/L
Endpoint(s) Effected: no effects on biomass and growth rate

Description of key information

In a 96-hour acute toxicity study, the cultures of the green alga, Selenastrum capricornutum, were exposed to spirodiclofen technical at the nominal concentrations of 20, 40 and 60 µg/L. The study included a control. The measured concentrations were 19.8, 41.5 and 61.6 µg/L. The test was a static design. The NOEC was 60 µg/L. The 96-hour EC50 based on growth rate and biomass was greater than 60 µg/L. This was the highest level tested since the spirodiclofen is insoluble at higher concentrations. There was no inhibition in the treatment levels as compared to the controls for growth rate or biomass.
No abnormalities were noted. This toxicity study is classified as acceptable and satisfies the guideline requirements for the aquatic nonvascular plant toxicity study with the green alga, Selenastrum capricornutum.
Results Synopsis
Test Organism: green alga (Pseudokirchneriella subcapitata or Selenastrum capricornutum)
Test Type: Static
96-hr EC50 >60 µg/L
96-hr NOEC >= 60 µg/L
Endpoint(s) Effected: no effects on biomass and growth rate

Key value for chemical safety assessment

EC50 for freshwater algae:

60 µg/L

EC10 or NOEC for freshwater algae:

60 µg/L