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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed according to relevant OECD guideline and compliant to GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
no
Details on sampling:
Due to the low solubility of the test item in polar and non polar solvents the determination of the test item concentration was not possible.

Test solutions

Vehicle:
no
Details on test solutions:
50 mg test substance was weighed into a 250 mL beaker, diluted with algal medium, transferred to a 500 mL Erlenmeyer flask and filled up to a volume of 500 mL. After treatment with ultrasonic waves for appr. 30 min the preparation was stirred for appr. 4 days and filtered through membrane filter (0.45 µm).

Test organisms

Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Strain: Desmodesmus subnspicatus CHODAT SAG 86.81
- Source (laboratory, culture collection): Sammlung von Algenkulturen (SAG), Pflanzenphysiologisches Institut der Universität Göttingen, D-37073 Göttingen
- Age of inoculum (at test initiation): four day-old preculture
- Method of cultivation: Fresh stocks were prepared every month on Z-Ager. Light intensity amounted 35-70 µE/m2 x s for 24 h per day

ACCLIMATION
- Acclimation period: four days, see above
- Culturing media and conditions (same as test or not): preculture incubated in 500 mL Erlenmeyer flasks with test medium; all algae were from the same source.
- Any deformed or abnormal cells observed: no

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Post exposure observation period:
None. Observations were carried out during exposure at the beginning of the experiment and after 24, 48 and 72 hours.

Test conditions

Test temperature:
22.2 - 24.2 °C (mean 23.2°C)
pH:
control 8.21 - 8.58
Nominal and measured concentrations:
A saturated solution prepared with 100 mg/L (nominal) was tested as limit concentration.
Details on test conditions:
TEST SYSTEM
- Test vessel: Sterile erlenmeyer flasks, volume: 250 mL
- Test volume: 100 mL
- Initial cells density: appr. 10000 cells/mL
- Control end cells density: 407190 (mean from 6 replicates) cells/mL
- No. of vessels per limit concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: dilution water according to guidelines
- Intervals of water quality measurement: at the start and at the end (72 h)

OTHER TEST CONDITIONS
- Photoperiod: 24 h/d light
- Light intensity and quality: nominally 60 - 120 μE·m-2·s-1

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Chlorophyll-impulsfluorometer
- Chlorophyll measurement: Chlorophyll-a-fluorescence, excitation at 435 nm, emission at 685 nm
- Other: measurements at the beginning of the test (after application) and every 24 h, filtrated culture medium was used as ground signal. Each replicate was measured 6-fold.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: only one concentration (saturated solution), limit test
- Justification for using less concentrations than requested by guideline: limit test
Reference substance (positive control):
yes
Remarks:
Potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 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
- Observation of abnormalities (for algal test): no
Results with reference substance (positive control):
- Results with reference substance valid? yes
- EC50 (biomass growth): 0.69 mg/l (0-72 h), 95 % confidence interval: 0.62-0.77 mg/l
- EC50 (rate-related inhibition): 0.89 mg/l (0-72 h), 95 % confidence interval: 0.73-1.09 mg/l
Reported statistics and error estimates:
The NOEC and LOEC were determined by calculation of statistical significance of biomass integrals and growth rates. One Way Analysis of Variance (ANOVA) and DUNNETT’s test was carried out for the determination of statistically significant differences compared to control replicates. When running a One Way Analysis of Variance a Normality Test and an Equal Variance Test were done first. P-values for both Normality and Equal Variance Tests were 0.05. The α-value for ANOVA was α=0.05.

Any other information on results incl. tables

Table 1: cell densities

Test concentration

Replicate No.

Cell density [cells/mL]

0 hours

Cell density [cells/mL]

24 hours

Cell density [cells/mL]

48 hours

Cell density [cells/mL]

72 hours

100 mg/L

(nominal)

1

9650

21920

127823

386348

2

9661

22084

124707

415561

3

10050

21612

139918

388501

Mean

9787

21872

130816

396803

Control

1

9384

20997

115338

380608

2

9599

22883

98672

423146

3

9784

22166

113309

419456

4

9579

22422

114457

405516

5

9517

20290

100763

428066

6

10153

20433

96396

386348

Mean

9669

21532

106489

407190

Table 2: evaluation after 72 h

Test concentration

Replicate No.

Biomass

Integral

Inhibition of Biomass

[%]

Growth Rate

Rate-related Inhibition

[%]

100 mg/L

(nominal)

1

318792

- 3.96

1.23

1.33

2

330419

- 7.47

1.25

- 0.57

3

330656

- 7.55

1.22

 2.29

Mean

(+)326622

- 6.24

(-)1.59

 1.02

Control

1

303179

1.23

2

309131

1.26

3

320743

1.25

4

315690

1.25

5

311294

1.27

6

284621

1.21

Mean

307444

1.25

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The test substance did not inhibit the biomass growth and rate related growth of green algae at the saturated nominal concentration of 100 mg/L.
Executive summary:

The toxicity of Pigment Red 254 to the unicellular freshwater green alga Desmodesmus subspicatus was determined according to the principles of OECD-Guideline 201 (1984) at DR.U.NOACK-LABORATORIEN in 31157 Sarstedt, Germany. The aim of the study was to assess the effects on growth rate and biomass production over a period of 72 h. The study was conducted under static conditions with an initial cell density of appr. 10000 cells/mL. A limit test with a saturated solution was carried out. A dispersion of 100 mg/L was prepared in dilution water and membrane filtrated with 0.45µm. Three replicates were tested for the saturated solution and 6 replicates for the control. Environmental conditions, pH-value and room temperature, were determined to be within the acceptable limits. All effect values are based on the saturated solution of the test item.

In this study Pigment Red 254. did not cause any inhibiting effects to Desmodesmus subspicatus after 72 h when tested with a saturated solution.