1-[(4-chlorophenyl)methyl]-1-cyclopentyl-3-phenylurea

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

Study period:

2000-01-12 to 2001-03-27

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

1984

Deviations:

no

Qualifier:

according to guideline

Guideline:

American Society for Testing and Materials. ASTM E 1218-04. Standard guide for conducting 96-h toxicity tests with microalgae. In: Annual Book of ASTM Standards, Vol. 11.06, West Conshohocken, PA. Current edition approved 2004

Version / remarks:

1990

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1989

Deviations:

no

Principles of method if other than guideline:

No major deviations except a slightly lower lux number (5992 lux instead of the recommended minimum of 6400 lux) which is not susceptible to influence the growth of Scenedesmus subspicatus negatively.

GLP compliance:

yes

Specific details on test material used for the study:

The test material is in a colorless powder form.

Analytical monitoring:

yes

Details on sampling:

Quantitative analyses of Pencycuron in samples of the nutrient medium were made at the beginning and the end of the experiment. The water samples were analysed by HPLC.

Vehicle:

yes

Remarks:

Dimethylformamide

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Controls: control and solvent control
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Dimethylformamide
- Evidence of undissolved material (e.g. precipitate, surface film, etc.):
- Other relevant information: A stock solution of the test item was prepared by adding 1.0001 g of Pencycuron techn. to a 5 mL volumetric flask and bringing this up to volume with Dimethylformamide. From this stock solution dilution series were made to reach the other concentrations. From these 0.1 mL were added to each test vessel of 1.0 L which were agitated with a magnetic stirrer for 30 minutes.

Test cultures and the cell-free, culture media used for quantitative analyses were prepared by mixing the appropriate quantities of the following components in the following order: 1) sterile, deionized water 2) 10-fold concentrated, nutrient solution, and 3) suitable volumes of the stock solution of the test item.

After mixing, the medium was divided into 2 parts. One part was used for the growth inhibition tests by inoculating it with enough 3-day old pre-culture to give a density of 1 x 10E4 algal cells/mL. This medium was divided into 150 mL aliquots which were poured into 300 mL Erlenmeyer flasks that were labelled with the study number, series number and concentration of test item. The flasks were sealed with cotton wool and/or cellulose plugs and placed in an incubator. A second part, used for quantitative analyses, was mixed with an appropriate amount of sterile, deionized water (instead of algal pre-culture) and incubated in the same way. All operations were carried out under sterile conditions.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green alga, Scenedesmus subspicatus
- Strain: SAG 86.81 ESP
- Method of cultivation: Stock cultures of the alga were grown at 23 ± 2°C under 16 h light/day in cotton and/or cellulose plugged, 300-mL Erlenmeyer flasks containing 50 mL of nutrient solution number 1. Fresh stock cultures were prepared once a week. All operations were conducted under sterile conditions.
Pre-cultures of the algae were inoculated with 1 x 10E4 cells/mL. These were grown in 200 mL nutrient solution number 2 for 3 days in an incubator, and then used to prepare treated and control cultures for the range finding- and final growth inhibition tests.

Test type:

static

Water media type:

other: Two solutions were used: Nutrient solution 1, which was used to grow stock cultures of algae, and Nutrient solution 2, which was used in all other tests with the algae.

Limit test:

no

Total exposure duration:

72 h

Test temperature:

24.0 - 24.7°C

pH:

Test start: 8.07 - 8.32 ; test end: 8.30 - 8.69

Nominal and measured concentrations:

Nominal concentrations of 0.10, 0.50, 1.00, 5.00, 10.0 and 20.0 mg a.s./L
Measured concentrations (day 0): 0.103, 0.491, 0.991, 2.92, 4.06, and 5.54 mg a.s./L
Measured concentrations (day 3): 0.109, 0.488, 0.823, 1.78, 2.51, and 17.1 mg a.s./L

Details on test conditions:

TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks
- Initial cells density: 1 x 10E4 algal cells/mL
- No. of vessels per concentration (replicates):3 replicates
- No. of vessels per control (replicates): 6 replicates each for the control and solvent control.

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Photoperiod: 16 h light/day
- Light intensity and quality: The incubator was illuminated with 5992 lux instead of the recommended minimum of 6400 lux which is not susceptible to influence the growth of algae negatively.

EFFECT PARAMETERS MEASURED
Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]: Cell numbers were estimated photometrically. For this purpose, samples of treated, inoculated culture medium were placed in 5 cm cuvettes and the extinctions were determined at a wave length of 578 nm using a single-beam-photometer (WTW MPM 1500). The photometer was calibrated using the treated culture medium without algae.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 0.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: based on test substance aqueous solubility

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 0.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: based on test substance aqueous solubility

Details on results:

The mean measured concentrations of the test substance varied between 28% and 103% of the nominal value at day 0 and between 25-109% at day 3. The recovery percentage decreased with increasing test concentrations. This indicates that the limit of water solubility (0.3 mg a.s./L) was reached, which was supported by the visual observation of un-dissolved material in the two highest test concentrations.

The pH values varied between 8.07 and 8.69 and temperatures between 23.5 and 25.4°C. The incubator was illuminated with 5992 lux instead of 8000 lux as given by the OECD 201 guideline.

A recovery percentage <80% was measured in fresh and aged test medium when the test concentration exceeds 1.0 mg/L and in the report, it was therefore concluded that for the parameter growth rate the 72h-ErC50 and NOEC are > 1.0 mg/L and =1.0 mg/L, respectively.

The illumination flux was lower than prescribed by the guidelines. As growth of the (control) cells was sufficient this is not considered to have influenced the results of the test.

All tested concentrations, except one, were above the water solubility of the test substance (0.3 mg a.s./L). It must be assumed that the amount of test substance dosed above the aqueous solubility of the active ingredient is not bio-available and does not contribute to the exposure of algae to the test substance. As no adverse effects to the algae were observed it can be concluded that under the circumstances of the test, the test substance is not toxic within its aqueous solubility.

Results with reference substance (positive control):

The concentrations tested were 0.10, 0.18, 0.32, 0.56, 1.00 and 1.80 mg a.s./L. In this test, the 0-72 hour ErC50determined for the biomass was 1.12 mg/L. The 0 to 72 hour average ErC50 determined for algal growth rate was 2.34 mg/L.

See "Attachments" in "Overall remarks, attachments" for the figure and tables.

Validity criteria fulfilled:

yes

Conclusions:

All tested concentrations, except one, were above the water solubility of the test substance (0.3 mg a.s./L). It must be assumed that the amount of test substance dosed above the aqueous solubility of the active ingredient is not bio-available and does not contribute to the exposure of algae to the test substance. As no adverse effects to the algae were observed it can be concluded that under the circumstances of the test, the test substance is not toxic within its aqueous solubility. Therefore, the the 72h-ErC50 is >0.3 mg a.s./L.

Executive summary:

The acute toxicity of Pencycuron Technical to Scenedesmus subspicatus was determined under static conditions.  Algae were exposed to nominal concentrations of 0.10, 0.50, 1.00, 5.00, 10.0 and 20.0 mg a.s./L, alongside a control and a solvent control.  The test substance is not toxic to Scenedesmus subspicatus at a level of its aqueous solubility, and the 72h-E_rC₅₀ is >0.3 mg a.s./L.

Description of key information

A study was conducted to determine the effect of Pencycuron on exponentially growing Selenastrum capricornutum over 72 hours under static exposure. Based on nominal concentrations of Pencycuron the 72-h E_rC₅₀ and NOEC (base on growth rate) were determined to be >0.3 and ≥0.3mg/L, respectively.

 

Test species	Result	Assessment	Reference
Scenedesmus subspicatus	72-h ErC50 > 0.3 mg a.s./L (nominal) 72-h NOErC ≥0.3 mg a.s./L (nominal)	Key study	Dorgerloh & Sommer (2001)

Key value for chemical safety assessment

EC50 for freshwater algae:

0.3 mg/L

EC10 or NOEC for freshwater algae:

0.3 mg/L