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
Study period:
06 August 1985 to 10 August 1985
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
According to the analytical results it can not be assured if a sufficient amount of test substance was used within the test.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985

Materials and methods

Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
-Effect concentration based on % water soluble fraction (WSF)
GLP compliance:
yes
Remarks:
EPA good Laboratory Practice Regulations set forth in 40 CFR Part 792.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Batch I


STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Stability under test conditions: The stability, identity, strength, and composition or other characteristics which appropriately identify the test substance are the responsibility of the Sponsor.


FORM AS APPLIED IN THE TEST (if different from that of starting material): A colorless liquid (not different from starting material)

OTHER SPECIFICS: The methods of synthesis, fabrication, and/or derivation of the test substance are documented and are the responsibility of the Sponsor.

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: Control, 6.25, 12.5, 25, 50 and 100%
- Sampling method: Samples for test material concentration verification were taken from the AAP nutrient media, prior to test material preparation, from all test treatments prior to adding test organisms, and from all treatments at study termination.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The MRD-84-521 water soluble fraction was prepared by combining 6.7 mL of test material with 1 liter of AAP nutrient media in a 2 liter flask. This mixture was slowly stirred for approximately 72 hours. After the mixing period, the mixture was
transferred to a separatory funnel and allowed to settle for approximately 1 hour. After the settling period, the water soluble fraction was drawn off and used as the 100 % stock solution.

Test organisms

Test organisms (species):
Scenedesmus capricornutum
Details on test organisms:
TEST ORGANISM
- Common name: Scenedesmus capricornutum
- Strain: Initial strain 1648
- Source (laboratory, culture collection): Department of Botany, University of Texas.
- Age of inoculum (at test initiation): The study was started with algal cells in the log phase of growth as calculated from growth of stock cultures. This is documented in the Algal Culture Log.
- Method of cultivation: The algae are cultured in AAP nutrient media prepared with distilled water and reagent grade chemicals. The media was prepared as per standard laboratory procedures. Culture vessels were sterile glass flasks whose volume
was approximately 60 % of capacity. The cultures were maintained in 400 (+/- 10%) footcandles of continuous cool white light, on a stirring platform with aeration, at a continuous temperature of 21 (+/- 1) degrees C. Stock cultures were maintained for approximately 14 days with new stock prepared weekly. Additional culture information is available in the Algal Culture Log at EHSL.

ACCLIMATION
- Culturing media and conditions (same as test or not): Same as test

Study design

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

Test conditions

Test temperature:
23.89 - 23.91
pH:
7.5 (+/- 0.1)
Nominal and measured concentrations:
Nominal: Control, 6.25, 12.5, 25, 50, 100 (% WAF)
Measured: -, 0.058, 0.219, 0.492, 0.873 (ppm total carbon)
Details on test conditions:
TEST SYSTEM
- Material, size, headspace, fill volume: size: 125 mL, fill volume: 50 mL
- Initial cells density: 2 X 10^4 cells/mL.
- Control end cells density: mean: 5.2 x 10^6 cells/mL.
- No. of vessels per concentration (replicates): 3 replicates
- No. of vessels per control (replicates): 3 replicates

GROWTH MEDIUM
- Standard medium used: yes (Miller, et al., 1978. The Selenastrum capricornutum Printz Algal Assay Bottle Test. U.S. EPA pp.17-18.)

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: The pH was adjusted where necessary to 7.5 (+/- 0.1) prior to adding test organisms.
- Photoperiod: 24 hours
- Light intensity and quality: 400 footcandles.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: The fluorescence readings were converted to cell numbers using a regression formula developed through cell counts (hemocytometer).
- Chlorophyll measurement: 24, 48, 72, and 96 hours

TEST CONCENTRATIONS
- Range finding study: no
Reference substance (positive control):
no

Results and discussion

Effect concentrations
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
not specified
Conc. based on:
test mat. (dissolved fraction)
Basis for effect:
growth rate
Remarks on result:
other: results statistically not evaluable
Details on results:
- Exponential growth in the control (for algal test): yes

Any other information on results incl. tables

No EC0 or EC50 values can be reported as there was insufficient growth inhibition in all of the test treatments. An earlier trial of this study was performed, however several of the treatments exhibited an unacceptable degree of variability in growth. In the past, this variability has been associated with contaminated glassware. The measured concentration of the test material in solution during the study is described by the Day 0 values only. The Total Carbon measurement is very sensitive to changes in carbonate and CO2 in solution. As the solution Total Carbon on Day 4 is the result of algal metabolism during the study, it is not a true representation of the test material in solution.

Table 1: Mean Cell Concentrations (cells/mL)

Date

Control

6.25

12.5

25.0

50.0

100.0

06-AUG-85

2.5 x 104

2.6 x 104

2.7 x 104

2.6 x 104

2.4 x 104

2.4 x 104

07-AUG-85

8.2 x 104

8.0 x 104

8.9 x 104

8.2 x 104

8.5 x 104

8.6 x 104

08-AUG-85

4.8 x 105

5.0 x 105

5.3 x 105

4.8 x 105

4.9 x 105

5.7 x 105

09-AUG-85

2.5 x 106

2.6 x 106

2.6 x 106

2.2 x 106

2.3 x 106

2.4 x 106

10-AUG-85

5.2 x 106

4.4 x 106

4.6 x 106

4.1 x 106

5.3x 106

5.2 x 106

 

Table 2: Total Carbon Content (ppm)

Sample

06-Aug-85

Day 0

(unfiltered)

10-Aug-85

Day 4

(filtered)

Control

3.374

10.01

6.25%

3.282

9.452

12.5%

3.432

9.829

25%

3.593

9.648

50%

3.866

9.195

100%

4.247

9.513

Applicant's summary and conclusion

Validity criteria fulfilled:
not specified
Conclusions:
Based on the estimated 96-h ErC50 of >100 mg/L, the test material is not classified as acutely toxic to freshwater algae according to Regulation (EC) No. 1272/2008 (CLP).
Executive summary:

No ErC0 or ErC50 values can be reported as there was insufficient growth inhibition in all of the test treatments after 96 hours. Therefore a 96 -h EC50 value of > 100 mg/L was estimated.