3-(difluoromethyl)-1-methyl-1H-pyrazole-4-carboxylic acid

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study with analytical monitoring

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0 (control), 120 mg/L as nominal concentrations
- Sampling method: Sampling was performed at the start of the exposure (0 h) from the inoculated replicate 7 of each concentration and the control. At the end of the exposure (72 h) samples were taken of the inoculated replicates 1-6 of the test concentrations and of the inoculated replicates 1-6 of the control group. The replicates were combined before sampling. Additionally, samples were collected from the uninoculated replicate 0 of each test concentration and the control at the end of the exposure (72 h).
- Sample storage conditions before analysis: Samples were transported to the Analytical Laboratory on the day of sampling. Ambient temperature

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The stock test solution (133.2 mg/L) was prepared by weighing 266.36 mg test substance and diluting it in 2 L test water. The mixture was stirred vigorously for about 10 minutes on a magnetic stir plate. To reach the correct nominal concentration (120 mg/L) at test start, 10 mL of inoculum culture was added to 90 mL of the stock test solution. The pH value of the stock solution was initially 4.3 and was adjusted with 1 molar NaOH to pH 8.4 prior to addition of algal inoculum.
The stock solution was visibly clear and slightly pink following preparation.
- Controls: The analytically determined concentrations of the test substance in the test solutions during the exposure period were within the range of + 20% of the nominal concentrations.

Test organisms

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: KORSHIKOV SAG 61.81
- Source (laboratory, culture collection): The algae were obtained at regular intervals from SAG (Collection of algal cultures in Göttingen) and were kept in liquid culture in the Laboratory of Ecotoxicology of the Experimental Toxicology and Ecology, BASF SE, Ludwigshafen, Germany.
- Age (on study day 0): A stock algal culture is maintained continuously at the test facility. Before the exposure an inoculum culture is prepared from the stock culture and incubated for 3 days at 21 – 24 °C (max. temperature difference 2 °C). After this time, the inoculum culture is in exponential growth phase and can be used to initiate the test (study day 0).

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Test temperature:

22.5 – 22.7°C

pH:

7.8-8.6

Nominal and measured concentrations:

0 (control), 120 mg/L as nominal concentrations

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks
- Type (delete if not applicable): closed, plugged with gas permeable silicone sponge caps
- Material, size, headspace, fill volume: glas, nominal volume 250 mL, fill volume: 100 mL
- Initial cells density: 5000 cells/mL
- No. of vessels per concentration (replicates): 6 replicates for each test substance concentration
- No. of vessels per control (replicates): 6 replicates for the Control
- 1 additional replicate per test group for initial concentration control analysis only.
- 1 additional uninoculated (without algae) replicate per test group for background fluorescence correction and to determine the influence of the test design on test substance stability.

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The test medium was prepared according to OECD Guideline for Testing of Chemicals, No. 201 Algal growth inhibition test, Mar 2006. The pH value of the test medium was 8.1.

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: 7.8-8.6
- Light intensity and quality: Average 4900 lux (within ± 15% variability) at a wave length of 400 - 700 nm
- Shaking rate: Continuous (approx. 85 rpm)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter] Test chamber: Vötsch Industrietechnik GmbH Bioline (VB1014) controlled climate cabinet.
- Chlorophyll measurement: Algal growth measured as in vivo chlorophyll-a fluorescence (pulsed excitation with light flashes having a wavelength of 432 nm)

TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 0 (control), 120 mg/L as nominal concentrations

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7)

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: algal yield

Effect concentrations 2

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: algal yield

Effect concentrations 3

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: algal yield

Effect concentrations 4

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 5

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 6

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- The cell multiplication factor in the untreated control was 198-fold after 72 hours. (counted cell density: 99 0000 cells/mL)
- Observation of abnormalities (for algal test): no remarkable observations

Results with reference substance (positive control):

- Results with reference substance valid? yes
- EC50: The ErC50 (72 h) of the control substance potassium dichromate was 0.97 mg/L (Date of the last control experiment: 29 Mar 2010 , project number: 60E0063/043031). These results indicate that the algae are responding normally to toxicant stress.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes