Registration Dossier

Ecotoxicological information

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:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Charge DEBR008539
Purity: 99.76% (GC-a/a), p-chlorobenzotrichloride < 0.1%
Specific details on test material used for the study:
Purity: OCTC 99.66%, PCTC 0.11% unknown 0.23%

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
The concentrations of o-chlorobenzoic acid in all test solutions and test cultures will be
measured at the beginning of exposure and 72 hours after the beginning of exposure by high­
performance liquid chromatography (HPLC). Samples taken from middle layer of vessels

Test solutions

Vehicle:
no
Details on test solutions:
The test substance is quickly hydrolyzed in water to produce o-chlorobenzoic acid as a
hydrolysate, and the half life of the test substance is less than 30 minutes (156.57 g of
o-chlorobenzoic acid is generated from 229.93 g of the test substance, calculated by the
testing facility). Therefore, the exposure test will be conducted after changing the test
substance completely to o-chlorobenzoic acid in test water.
The test concentration will be expressed by both the test substance and o-chlorobenzoic
acid. The concentration of o-chlorobenzoic acid will be measured in test solutions and test
culutures, and the effect concentration will be calculated based on the concentration of
o-chlorobenzoic acid. The obtained toxicity data will be converted into the concentration of
the test substance.
Stock volumes of 100mg/L are prepared using dilution water and stirred for 48 hours at 22C.

Test organisms

Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: Unicellular green algae
- Strain: ATCC22662
- Source (laboratory, culture collection): ATCC
- Age of inoculum (at test initiation): 3-4 days
- Method of cultivation: period subculture in Gorham medium

ACCLIMATION
- Acclimation period: 3-4 days
- Culturing media and conditions (same as test or not): as test
- Any deformed or abnormal cells observed:

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h

Test conditions

Test temperature:
22 +/- 2C
pH:
8.0( 7.7 to 8.1 at 22 C)
Nominal and measured concentrations:
Nominal concentration of OCTC: 0, 10, 18, 32, 56, 100 mg/L
Nominal concentration of o-chlorobenzoic acid: 0, 6.8, 12, 22, 38, 68 mg/L
measured concentration of o-chlorobenzoic acid 0 hr: 6.56,11.8,20.4, 35.7, 63.4 mg/L
measured concentration of o-chlorobenzoic acid 72 hrs: 6.28, 11.3, 20.3, 34.7, 60.5 mg/L
Mean measured concentration of o-chlorobenzoic acid: 6.42, 11.5, 20.3, 35.2, 61.9 mg/L

Details on test conditions:
TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: glass 300 ml fill volume
- Aeration: cultures shaken 100 rpm
- Initial cells density: 5x10^3 cells/mL
- Control end cells density:
- No. of organisms per vessel:
- No. of vessels per concentration (replicates): 3 (three)
- No. of vessels per control (replicates): 6 (six)
- No. of vessels per vehicle control (replicates): 6 (six)

GROWTH MEDIUM
- Standard medium used: yes
- Detailed composition if non-standard medium was used:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides:
- Chlorine:
- Alkalinity:
- Ca/mg ratio:
- Conductivity:
- Culture medium different from test medium:
- Intervals of water quality measurement:

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: 65-75 µE/m^2/s white fluorescent lamp
- Salinity (for marine algae):

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]
- Chlorophyll measurement:
- Other:

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Justification for using less concentrations than requested by guideline: base on results of rangefinding study
- Range finding study
- Test concentrations: 0.1, 1, 10 ,100 mg/L
- Results used to determine the conditions for the definitive study: yes
Reference substance (positive control):
yes
Remarks:
potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
35.1 mg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
16.9 mg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
test mat.
Basis for effect:
growth rate

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The ErC50 of the test substance was found to be 35.1 mg/L and the NOEC was 16.9 mg/L