Chromate(5-), bis[4-(hydroxy-κO)-7-[2-(2-hydroxy-1-naphthalenyl)diazenyl]-3-[2-[2-(hydroxy-κO)- 3-nitro-5-sulfophenyl]diazenyl]-2-naphthalenesulfonato(4-)]-, sodium (1:5)

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

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

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Analytical monitoring:

no

Details on sampling:

During the final test duplicate samples were taken from 1.0, 3.2 and 10 mg/l and the blank control.
-Frequency: at t=0 h and t=72 h.
-Volume: 25 ml
-Storage: Samples were transferred to Analytical Chemistry directly after sampling.
Additionally, reserve samples of 25 ml were taken from all test solutions and stored in a deep freezer until possible analysis for a maximum of three months or until delivery of the final report.
-Storage: All samples not analysed on the same day of sampllng were frozen until analysis.
-Sample pretreatment: The frozen samples were defrosted at room temperature. The samples of t=72 hours (except for the sample of 10 mg/l without algae) were centrifuged for 5 minutes at 2000 rpm. The samples of 0 and 1 mg/l were not diluted. The entire volume of the samples of 3.2 and 10 mg/l was transferred quantitatively into a sultable volumetric flask. The flask was filled up to the mark with Milli-Q water.

Vehicle:

no

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
-Strain: 86.81 SAG.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

21 - 21.5 °C

pH:

8.6 - 8.2

Nominal and measured concentrations:

Nominal: 1.0, 1.8, 3.2, 5.6 and 10 mg/l
Analysis of the samples taken at the start (t=0) of the final test revealed mean actual concentrations of 0.90 mg/l (90 %), 2.86 mg/l (89 %) and 9.21 mg/l (92 %) at the nominal concentrations of 1.0, 3.2 and 10 mg/l, respectively.

Details on test conditions:

A range-finding test preceded the final test to provide information about the range of concentrations to be used in the final test. The range to which algae were exposed was 0.1 to 100 mg/l, increasing by a factor 10.

TEST SYSTEM
-Test vessel: 100 ml, all-glass
-Cells density: 1 x 10^4 cells/ml using a 4 days old preculture with a density of 195 x 10^4 cells/ml
-Replicas: 3 replicas of each test concentration, 6 replicas of the blank-control,1 replica of each test concentration without algae
-Incubation: during incubation the algal cells were kept in suspension by continuous shaking.

GROWTH MEDIUM
-Standard medium used: yes

TEST MEDIUM
The test media were prepared using a stock solution in ISO-medium. For this purpose the test substance was added quantitatively to obtain a nominal concentration of 100 mg/l. From this stock solution the required volumes for preparation of the test concentrations were added to ISO-medium. The final volume of the test solutions was 50 ml per vessel. At the start of the test the test solutions ranged from light to clear blue. One extra replica without algae was included for each concentration to correct for background absorbance.

ISO-medium formulated according to the International Standards "Algal growth inhibition test" November, 1989; formulated using Milli-Q water in such a way that precipitation does not occur.
NH4CI 15 mg/l
MgCl2.6H20 12 mg/l
CaCl2.2H20 18 mg/l
MgS04.7H20 15 mg/l
KH2PO4 1.6 mg/l
FeCl3.6H20 80 µg/l
Na2EDTA.2H20 100 µg/l
H3BO3 185 µg/l
MnCl2.4H20 415 µg/l
ZnCl2 3 µg/l
ZnCl2.6H20 1.5 µg/l
ZnCl2.2H20 0.01 µg/l
Na2MoO4.2H20 7 µg/l
NaHCO3 50 mg/l

OTHER TEST CONDITIONS
-Photoperiod: continuously
-Light intensity and quality: TLD-lamps of 18 Watt (Philips. Spain) yielding 6500-7000 lux.

STABILITY TEST
The substance was determined to be stable under test conditions for 72 hours.

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 10 mg/L

Nominal / measured:

estimated

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

algicidal effect

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

7.34 mg/L

Nominal / measured:

estimated

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

algistatic effect

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

2.67 mg/L

Nominal / measured:

estimated

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.81 mg/L

Nominal / measured:

estimated

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: Since there was no NOEC for cell growth inhibition, the EC10 for growth inhibition was calculated

Details on results:

-At the end of the exposure period, the actual concentrations were 106 and 109 % of nominal measured at 1.0 and 3.2 mg/l, respectively. The actual concentration remained constant at 10 mg/l.
-Range-finding: significant growth inhibition was recorded at 10 and 100 mg/l.
-Inhibition of cell growth increased with increasing concentration of the substance resulting in 92 % inhibition at 10 mg/l. Statistically significant inhibition of cell growth was found at all test concentrations (Williams' test: P=0.05). Growth rate reduction increased with increasing concentration of the substance resulting in 59 % reduction at 10 mg/l. Statistically significant reduction of growth rate was found at test concentrations of 1.8 mg/l and higher (Williams' test: P=0.05). The additional growth test, however, showed total recovery of cell growth in the algal suspensions originating from the test concentration of 10 mg/l. Hence, the effects on cell growth recorded at 10 mg/l (and lower) were not due to a toxic effect but probably caused by the interference of the coloured solutions with the wavelengths required for normal algal functioning (algistatic effect).
-Acceptability of the test: in the controls the cell density increased by an average factor of 45 within 3 days. Further, all test conditions remained within the ranges prescribed by the protocol.

Validity criteria fulfilled:

yes

Conclusions:

ErC50 (72 h) > 10 mg/l based on agicidal effect

Executive summary:

The acute toxicity of the test item to aquatic algae and cyanobacteria was investigated in a 96 -hour static test to Scenedesmus subspicatus, according to the OECD Guideline 201, the method C.3 of EEC-Directive 92/69 EEC and the ISO International Standard 8692. No significant reduction of growth rate was recorded at 1.0 mg/l (NOErC). As the test substance is coloured, the cell growth inhibition most likely was related interception of wavelengths necessary for normal algal functioning (algistatic effect). Although the test item induced an algistatic effect on cell growth at concentrations up to 10 mg/l, it did not induce an algicidal effect at 10 mg/l.

Thus, the ErC50 (72 h) for algicidal effect was found to be > 10 mg/l.

Description of key information

ErC50 (72 h) > 10 mg/l

Additional information

The acute toxicity of the test item to aquatic algae and cyanobacteria was investigated in a 96 -hour static test to Scenedesmus subspicatus,according to the OECD Guideline 201, the method C.3 of EEC-Directive 92/69 EEC and the ISO International Standard 8692.

The ECr50 (72 h) for algicidal effect was found to be greater than 10 mg/l.